Fungal protein disulfide isomerase

ABSTRACT

PCT No. PCT/DK94/00266 Sec. 371 Date Dec. 11, 1995 Sec. 102(e) Date Dec. 11, 1995 PCT Filed Jun. 28, 1994 PCT Pub. No. WO95/00636 PCT Pub. Date Jan. 5, 1995A fungal protein disulfide isomerase obtainable from fungal species belonging to Aspergillus, especially A. oryzae, or A. niger, is disclosed. Furthermore sequences for recombinant production of the protein disulfide isomerase are disclosed.

FIELD OF THE INVENTION

The present invention relates to an active recombinant fungal proteindisulfide isomerase, compositions comprising said fungal proteindisulfide isomerase, and methods for their use; a DNA constructcomprising a DNA sequence encoding said fungal protein disulfideisomerase, and a vector and cell harbouring the DNA construct.Furthermore, the present invention relates to a method of preparing thefungal protein disulfide isomerase by use of both traditional andrecombinant DNA techniques.

BACKGROUND OF THE INVENTION

The use of protein disulfide redox agents such as protein disulfideisomerases (PDI), and thioredoxins (TRX) for various purposes has beenknown for some time.

Protein disulfide redox agents catalyse the general reaction:

    R.sub.1 --SH+R.sub.2 --SH+Enz.sub.ox ⃡R.sub.1 --S--S--R.sub.2 +Enz.sub.red                                              (reaction I)

where R₁ and R₂ represent protein entities which are the same ordifferent, either within the same polypeptide or in two polypeptides,Enz_(ox) is a protein disulfide redox agent in the oxidised state, andEnz_(red) is a protein disulfide redox agent in the reduced state. EC5.3.4.1 refers to an enzyme capable of capable of catalysing therearrangement of --S--S-- bonds in proteins and EC 1.6.4.4 and EC1.8.4.2 is an example of enzymes catalysing the reaction with NAD(P)Hand glutathione as a mediator, respectively.

This type of activity has been designated as protein disulfideisomerase, sulfhydryl oxidase, protein disulfide reductase, disulfideisomerase, protein disulfide transhydrogenase, and sulfhydryl oxidase.

Disulfide linkages in proteins are formed between cysteine residues andhave the general function of stabilising the three dimensional structureof the proteins. They can be formed between cysteine residues of thesame or different polypeptides.

Disulfide linkages are present in many types of proteins such asenzymes, structural proteins, etc. Enzymes are catalytic proteins suchas proteases, amylases, etc., while structural proteins can bescleroproteins such as keratin, etc. Protein material in hair, wool,skin, leather, hides, food, fodder, stains, and human tissue containsdisulfide linkages. Treatment of some of these materials with PDI andTRX, and a redox partner have been described previously.

The use of TRX for waving, straightening, removing and softening ofhuman and animal hair was described by Pigiet et al. (EP 183506 and WO8906122). Pigiet (U.S. Pat. No. 4,771,036) also describes the use of TRXfor prevention and reversal of cataracts. Schreiber (DE 2141763 and DE2141764) describes the use of protein disulfide transhydrogenase forchanging the form of human hair. Pigiet (EP 225156) describes the use ofTRX for refolding denatured proteins. Use of TRX to prevent metalcatalysed oxidative damage in biological reactions is described byPigiet et al. (EP 237189).

Toyoshima et al. (EP 277563 and EP 293793) describes the use of PDI tocatalyse renaturation of proteins having reduced disulfide linkages orunnatural oxidised disulfide linkages, in particular in connection withrenaturation of recombinantly produced proteins. Brockway (EP 272781),and King and Brockway (EP 276547) describe the use of PDI forreconfiguration of human hair, and for treatment of wool, respectively.Sulfhydryl oxidase for the treatment of Ultra-high temperaturesterilized milk is described in U.S. Pat. No. 4,894,340, U.S. Pat. No.4,632,905, U.S. Pat. No. 4,081,328 and U.S. Pat. No. 4,053,644.Schreiber (DE 2141763 and DE 2141764) describes the use of proteindisulfide transhydrogenase for changing the form of human hair.

The uses of such enzymes have all been connected with reduction ofprotein disulfide linkages to free protein sulhydryl groups and/or theoxidation of protein sylfhydryl groups to protein disulfide linkages,and/or the rearrangement of disulfide linkages in the same or betweendifferent polypeptides, and sometimes to the use of these processes insequence.

Protein disulfide redox agents can be divided into two main groups ofenzymes, thioredoxin type (TRX), and protein disulfide isomerase type(PDI).

Both these can be modified to obtain protein engineered derivatives,chemical modifications and hybrids of TRX and/or PDI (ENG).

TRX is a 12-kDa protein having a redox-active disulfide/dithiol andcatalysing thiol-disulfide exchange reactions (Edman et al., Nature317:267-270, 1985; Holmgren, Ann. Rev. Biochem. 54:237-271, 1985;Holmgren, J. Biol. Chem. 264:13963-13966, 1989). PDI consists of twosubunits, each consisting of two domains which are homologous to TRX.

TRX and PDI can be obtained from a number of sources: PDI: proteindisulfide isomerases have mainly been identified from mammalian sources,such as Bovine (Yamauchi et al., Biochem. Biophys. Res. Commun.146:1485-1492, 1987), Chicken (Parkkonen et al., Biochem. J.256:1005-1011, 1988), Human (Rapilajaniemi et al. EMBO J. 6:643-649,1987), Mouse (Gong, et al., Nucleic Acids Res. 16:1203, 1988), Rabbit(Fliegel et al., J. Biol. Chem. 265:15496-15502, 1990), and Rat (Edmanet al., Nature 317:267-270, 1985). PDI has furthermore been isolatedfrom yeast (Tachikawa et al., J. Biochem. 110:306-313).

TRX: Thioredoxin has been identified from bacteriophages, bacteria suchas Escherichia coli (Wallace and Kusher, Gene 32:399-408, 1984) andBacillus subtilis (Chen et al. J. Biol Chem. 262:8787-8798, 1987) andeukaryotes.

It would be desirable to facilitate the production of protein disulfideisomerase (PDI), to be able of producing both larger amounts of theenzyme and to produce it in a more economical manner than what ispossible by the prior art methods.

Engineered variants (ENG) with improved properties for particularapplications are also highly desirable and can be prepared by a varietyof methods based on standard recombinant DNA technology:

1) by using site-directed or random mutagenesis to modify the genesencoding TRX or PDI in order to obtain ENG with one or few amino acidchanges,

2) by inhibiting or otherwise avoiding dimerisation of the subunits ofPDI, thus giving rise to PDI monomers,

3) by producing partial monomers of PDI or TRX, in which regions of theNH2-- or COOH termini of PDI or TRX are lacking,

4) by creating hybrids of PDI, TRX and/or ENG,

5) by chemically or enzymatically modifying the products of 1)-4),

6) by a combination of any of 1)-5).

ENG produced according to 1) were described by Lundstrom et al. (J.Biol. Chem. 267:9047-9052, 1992) and by a combination of 3) and 5) byPigiet (WO 8906122).

PDI, and TRX can, apart from their natural sources, be obtained byexpression of recombinant DNA encoding plant, animal, human or microbialPDI, or TRX, in various hosts, such as microorganisms followed bypurification of PDI, or TRX from extracts or supernatants of said hostorganisms. This goes also for ENG. Preparation of Trx from naturalsources is described by Luthman and Holmgren (Biochem. 121:6628-6633,1982), Wada and Buchanan (in "Thioredoxins, structure and function"(Gadal, Ed.) Editions du Centre National de la Recherche Scientifique),Porque et al. (J. Biol. Chem. 245:2362-2379, 1970) and by Laurent et al.(J. Biol. Chem. 239:3436-3445), whereas recombinant production of TRX isdescribed by Krause et al. (J. Biol. Chem. 266:9494-9500). PDI orsulfhydryl oxidase has been prepared from natural sources by Lambert andFreedman (Biochem J. 213:225-234, 1983), Starnes et al. (U.S. Pat. No.4,632,905) and Hammer et al. (U.S. Pat. No. 4,894,340), and byrecombinant technology by among others Yamauchi et al. (Biochem.Biophys. Res. Commun. 146:1485-1492, 1987). Finally, recombinantproduction of an ENG is described by Lundstrom et al. (J. Biol. Chem.267:9047-9052, 1992).

SUMMARY OF THE INVENTION

The present inventors have succeeded in cloning a DNA sequence encodinga fungal protein disulfide isomerase from filamentous fungi and inobtaining expression of an active protein disulfide isomerase from saidDNA sequence, both in the same species and in other organisms,especially microorganisms, and preferably in fungi.

Accordingly, in a first aspect the present invention relates to anactive protein disulfide isomerase obtainable from filamentous fungi,specifically fungi belonging to the genus Aspergillus, and especially aprotein disulfide isomerase obtainable from A. oryzae, or A. niger,which enzyme is immunologically reactive with an antibody raised againsta purified protein disulfide isomerase derived from Aspergillus oryzae,IFO 4177, or Aspergillus niger, A524.

From the sequence of the isolated enzyme it can be seen that the proteindisulfide isomerase has two -Cys-X-Y-Cys- subunits in positions 58-61and 393-396. The invention consequently also comprises active truncatedforms of the enzymes of the invention, wherein at least one subunit isretained. Examples hereof could be an enzyme having an amino acidsequence corresponding to the residues 20 to 100, residues 330 to 450,or residues 360 to 430 of the appended SEQ ID No. 3, or thecorresponding sequence of the enzyme of the invention in question.

Under this aspect, the invention specifically relates to enzymesexhibiting protein disulfide isomerase activity comprising the aminoacid residues 1-131, 1-141, 1-143, 1-163, 1-174, or 1-281, of the aminoacid sequence shown in the appended SEQ ID No. 3, or variants thereofexhibiting a protein disulfide isomerase activity. Further specificenzymes are enzymes exhibiting protein disulfide isomerase activitycomprising the amino acid residues 1-115, of the amino acid sequenceshown in the appended SEQ ID No. 3 extended with the following sequence:

Leu-Ile-Arg-Glu-Leu-Leu-Gln-Glu-Leu-Val-Asn-Lys-His-Leu (SEQ ID NO.11);and an enzyme comprising the amino acid residues 1-511, of the aminoacid sequence shown in the appended SEQ ID No. 3, and wherein the aminoacid residue in position 511 is changed from Glu to Ala.

In the present context, the term "derived from" is intended not only toindicate a protein disulfide isomerase produced by strains IFO 4177 orA524, but also a protein disulfide isomerase encoded by a DNA sequenceisolated from these strains such as indicated in SEQ ID No. 1 and SEQ IDNo. 2, or a sequence homologous thereto encoding a polypeptide withprotein disulfide isomerase activity and produced in a host organismtransformed with said DNA sequence.

Accordingly, the present invention thus relates to an enzyme exhibitingprotein disulfide isomerase activity, which enzyme is immunologicallyreactive with an antibody raised against a purified protein disulfideisomerase derived from Aspergillus oryzae, IFO 4177.

In the present context, the term "homologue" is intended to indicate apolypeptide encoded by DNA which hybridizes to the same probe as the DNAcoding for the protein disulfide isomerase enzyme under certainspecified conditions (such as presoaking in 5×SSC and prehybridizing for1 h at ˜40° C. in a solution of 5×SSC, 5×Denhardt's solution, 50 mMsodium phosphate, pH 6.8, and 50 μg of denatured sonicated calf thymusDNA, followed by hybridization in the same solution supplemented with 50μCi 32-P-dCTP labelled probe for 18 h at ˜40° C. followed by washingthree times in 2×SSC, 0.2% SDS at 40° C. for 30 minutes). Morespecifically, the term is intended to refer to a DNA sequence which isat least 70% homologous to the sequences indicated above encoding theprotein disulfide isomerase of the invention. The term is intended toinclude modifications of the DNA sequences indicated above, such asnucleotide substitutions which do not give rise to another amino acidsequence of the protein disulfide isomerase but which correspond to thecodon usage of the host organism into which the DNA construct isintroduced or nucleotide substitutions which do give rise to a differentamino acid sequence and therefore, possibly, a different proteinstructure which might give rise to a protein disulfide isomerase mutantwith different properties than the native enzyme. Other examples ofpossible modifications are insertion of one or more nucleotides into thesequence, addition of one or more nucleotides at either end of thesequence, or deletion of one or more nucleotides at either end or withinthe sequence.

In the present context the term active protein disulfide isomerase isintended to indicate an enzyme having an activity similar to that ofprotein disulfide isomerase, i.e. an enzyme capable of catalysingreaction I. The activity may be determined in an assay based onoxidative refolding of reduced Bowman-Birk soya bean inhibitor, e.g. asdescribed in the Materials and Methods section below.

The term "recombinant" as used about the protein disulfide isomerase ofthe invention is intended to indicate that it is produced by a celltransformed with a DNA sequence encoding the protein disulfideisomerase. Thus, the recombinant protein disulfide isomerase may beproduced by either its parent organism or another organism.

In a further aspect the present invention relates to a DNA constructcomprising a DNA sequence encoding an active recombinant proteindisulfide isomerase of the invention as defined above. Such a DNAconstruct may comprise introns (an example thereof is shown in theappended SEQ ID No. 1) and/or regulatory elements native to the partscoding for the mature protein disulfide isomerase of the invention, orbe a cDNA construct comprising only that part coding for the matureprotein disulfide isomerase (an example being the appended SEQ ID No.2).

In still further aspects the present invention relates to a recombinantexpression vector harbouring the DNA construct of the invention, to acell which either harbours the DNA construct or the expression vector ofthe invention, and to a process for the production of a proteindisulfide isomerase of the invention, wherein a cell of the invention asdescribed above is cultured under conditions conducive to the productionof the protein disulfide isomerase, and the protein disulfide isomeraseis subsequently recovered from the culture.

Finally, the present invention relates to compositions comprising theactive protein disulfide isomerase of the invention and methods fortheir use in various applications.

BRIEF DESCRIPTION OF THE TABLES AND DRAWING

The invention is further illustrated in the accompanying tables anddrawing, in which

Table 1 shows an alignment of published eukaryotic PDI amino acidsequences: Bovine (Bos taurus) (Yamauchi et al., Biochem. Biophys. Res.Commun. 146:1485-1492, 1987), chicken (Gallus gallus) (Parkkonen et al.,Biochem. J. 256:1005-1011, 1988), human (Homo sapiens) (Rapilajaniemi etal. EMBO J. 6:643-649, 1987), mouse (Mus musculus) (Gong, et al.,Nucleic Acids Res. 16:1203, 1988), rabbit (Oryctolagus cuniculus)(Fliegel et al., J. Biol. Chem. 265:15496-15502, 1990), rat (Rattusnorvegicus) (Edman et al., Nature 317:267-270, 1985), and yeast(Saccharomyces cerevisiae) (Tachikawa et al., J. Biochem. 110:306-313)(SEQ ID NOS:26-32, respectively).

Table 2 shows an alignment of PDI amino acid sequences: Alfalfa(Medicago sativa) (Shorrosh and Dixon, Plant. Mol. Bio. 19:319-321,1992), A. oryzae (this invention), yeast (Saccharomyces cerevisiae)(Tachikawa et al., J. Biochem. 110:306-313), bovine (Bos taurus)(Yamauchi et al., Biochem. Biophys. Res. Commun. 146:1485-1492, 1987),rat (Rattus norvegicus) (Edman et al., Nature 317:267-270, 1985), andmouse (Mus musculus) (Gong, et al., Nucleic Acids Res. 16:1203, 1988)(SEQ ID NOS:33-38, respectively), and

FIG. 1 illustrates the construction of the expression plasmids pCaHj431,pCaHj432, pCaHj433, and pCaHj434 further described in Example 1.

DETAILED DESCRIPTION OF THE INVENTION

The amino acid sequence of the protein disulfide isomerase of theinvention, which was isolated from a strain of the A. oryzae, has beenaligned with that of protein disulfide isomerases of other origins andhave been shown to have a degree of identity of about 38% with that ofSaccharomyces cerevisiae (GenBank Acc. No. M62815) and 30% with that ofAlfalfa (GenBank Acc. No. 11499).

These homologies are taken to indicate that some kind of evolutionaryrelationship exists between protein disulfide isomerases, and that theprotein disulfide isomerase of the invention may represent a distinctclass of protein disulfide isomerase. It is contemplated that theprotein disulfide isomerase of the invention or DNA encoding the proteindisulfide isomerase may be isolated from other organisms, includinganimals, especially a mammal, an insect, a plant or a microorgamism. Inthe present context, especially interesting origins are bacteria andfungi, including yeasts and filamentous fungi.

As indicated above the sequence of the isolated enzyme shows that theprotein disulfide isomerase of the invention has two -Cys-X-Y-Cys-subunits in positions 58-61 and 393-396.

The invention consequently also comprises active truncated forms of theenzymes of the invention, wherein at least one subunit is retained.Examples hereof could be an enzyme having an amino acid sequencecorresponding to the residues 20 to 100, residues 330 to 450, orresidues 360 to 430 of the appended SEQ ID No. 3, or the correspondingsequence of the enzyme of the invention in question.

Under this aspect, the invention specifically relates to enzymesexhibiting protein disulfide isomerase activity comprising the aminoacid residues 1-131 (SEQ ID No. 10), 1-141 (SEQ ID No.9), 1-143 (SEQ IDNo. 8), 1-163 (SEQ ID No. 7), 1-174 (SEQ ID No. 6), 1-281 (SEQ ID No.5), or 25-225 (SEQ ID No. 12) of the amino acid sequence shown in theappended SEQ ID No. 3, or variants/derivatives thereof exhibiting aprotein disulfide isomerase activity. Further specific enzymes areenzymes exhibiting protein disulfide isomerase activity comprising theamino acid residues 1-115, of the amino acid sequence shown in theappended SEQ ID No. 3 extended with the following sequence:

Leu-Ile-Arg-Glu-Leu-Leu-Gln-Glu-Leu-Val-Asn-Lys-His-Leu (SEQ ID No. 11);and an enzyme comprising the amino acid residues 1-511, of the aminoacid sequence shown in the appended SEQ ID No. 3, and wherein the aminoacid residue in position 511 is changed from Glu to Ala (SEQ ID No. 4).

The DNA sequence of the DNA construct of the invention encoding arecombinant protein disulfide isomerase enzyme as defined above ispreferably as shown in the appended SEQ ID No. 1 (genomic DNA) or SEQ IDNo. 2 (cDNA). Analogues of said sequences, which differ in one or morecodons, but which encodes the recombinant protein disulfide isomeraseare also within the invention.

Similar DNA sequences coding for the truncated forms of the proteindisulfide isomerases of the invention are also part of the invention.DNA sequences therefore can be taken from SEQ ID No. 1, or preferablySEQ ID No. 2.

The DNA sequence of the DNA construct of the invention may be isolatedby well-known methods. Thus, the DNA sequence may, for instance, beisolated by establishing a cDNA or genomic library from an organismexpected to harbour the sequence, and screening for positive clones byconventional procedures. Examples of such procedures are hybridizationto oligonucleotide probes synthesized on the basis of the full aminoacid sequence shown in SEQ ID No. 3, or a subsequence thereof inaccordance with standard techniques (cf. Sambrook et al., 1989), and/orselection for clones expressing a protein disulfide isomerase activityas defined above, and/or selection for clones producing a protein whichis reactive with an antibody raised against the protein disulfideisomerase comprising the amino acid sequence shown in SEQ ID No. 3 andin particular amino acid residues 1-143 thereof as shown in SEQ ID No.8.

A preferred method of isolating a DNA construct of the invention from acDNA or genomic library is by use of polymerase chain reaction (PCR)using degenerate oligonucleotide probes prepared on the basis of theamino acid sequence of the protein disulfide isomerase of the inventioncomprising amino acid residues 1-515 of SEQ ID No. 3. For instance, thePCR may be carried out using the techniques described in U.S. Pat. No.4,683,202 or by R. K. Saiki et al. (1988).

Alternatively, the DNA sequence of the DNA construct of the inventionmay be prepared synthetically by established standard methods, e.g. thephosphoamidite method described by Beaucage and Caruthers (1981), or themethod described by Matthes et al. (1984). According to thephosphoamidite method, oligonucleotides are synthesized, e.g. in anautomatic DNA synthesizer, purified, annealed, ligated and cloned inappropriate vectors.

Finally, the DNA construct may be of mixed genomic and synthetic, mixedsynthetic and cDNA or mixed genomic and cDNA origin prepared by ligatingfragments of synthetic, genomic or cDNA origin (as appropriate), thefragments corresponding to various parts of the entire recombinant DNAmolecule, in accordance with standard techniques.

DNA constructs coding for the truncated forms of the enzyme of theinvention may naturally be made in corresponding ways.

The recombinant expression vector carrying the DNA construct of theinvention may be any vector which may conveniently be subjected torecombinant DNA procedures, and the choice of vector will often dependon the host cell into which it is to be introduced. Thus, the vector maybe an autonomously replicating vector, i.e. a vector which exists as anextrachromosomal entity, the replication of which is independent ofchromosomal replication, e.g. a plasmid, a bacteriophage or anextrachromosomal element, minichromosome or an artificial chromosome.Alternatively, the vector may be one which, when introduced into a hostcell, is integrated into the host cell genome and replicated togetherwith the chromosome(s) into which it has been integrated.

In the vector, the DNA sequence should be operably connected to asuitable promoter sequence. The promoter may be any DNA sequence whichshows transcriptional activity in the host cell of choice and may bederived from genes encoding proteins either homologous or heterologousto the host cell. Examples of suitable promoters for directing thetranscription of the DNA construct of the invention, especially in abacterial host, are the promoter of the lac operon of E. coli, theStreptomyces coelicolor agarase gene dagA promoters, the promoters ofthe Bacillus licheniformis α-amylase gene (amyL), the promoters of theBacillus stearothermophilus maltogenic amylase gene (amyM), thepromoters of the Bacillus Amyloliquefaciens α-amylase (amyQ), thepromoters of the Bacillus subtilis xylA and xylB genes etc. Fortranscription in a fungal host, examples of useful promoters are thosederived from the gene encoding A. oryzae TAKA amylase, Rhizomucor mieheiaspartic proteinase, A. niger neutral α-amylase, A. niger acid stableα-amylase, A. niger glucoamylase, Rhizomucor miehei lipase, A. oryzaealkaline protease, A. oryzae triose phosphate isomerase or A. nidulansacetamidase.

The expression vector of the invention may also comprise a suitabletranscription terminator and, in eukaryotes, polyadenylation sequencesoperably connected to the DNA sequence encoding the recombinant proteindisulfide isomerase of the invention. Termination and polyadenylationsequences may suitably be derived from the same sources as the promoter.

The vector may further comprise a DNA sequence enabling the vector toreplicate in the host cell in question. Examples of such sequences arethe origins of replication of plasmids pUC19, pACYC177, pUB110, pE194,pAMB1 and pIJ702.

The vector may also comprise a selectable marker, e.g. a gene theproduct of which complements a defect in the host cell, such as the dalgenes from B. subtilis or B. licheniformis, or one which confersantibiotic resistance such as ampicillin, kanamycin, chloramphenicol ortetracyclin resistance. Examples of Aspergillus selection markersinclude amdS, argB, niaD and sC, a marker giving rise to hygromycinresistance. Furthermore, the selection may be accomplished byco-transformation, e.g. as described in WO 91/17243.

While intracellular expression may be advantageous in some respects,e.g. when using certain bacteria as host cells, it is generallypreferred that the expression is extracellular. The protein disulfideisomerase of the invention or truncated forms thereof comprising theamino acid sequences shown in the SEQ ID Nos. 3 to 12 may furthermorecomprise a preregion permitting secretion of the expressed proteindisulfide isomerase into the culture medium. If desirable, thispreregion may be native to the protein disulfide isomerase of theinvention or substituted with a different preregion or signal sequence,conveniently accomplished by substitution of the DNA sequences encodingthe respective preregions.

The procedures used to ligate the DNA construct of the invention, thepromoter, terminator and other elements, respectively, and to insertthem into suitable vectors containing the information necessary forreplication, are well known to persons skilled in the art (cf., forinstance, Sambrook et al. (1989)).

The cell of the invention either comprising a DNA construct or anexpression vector of the invention as defined above is advantageouslyused as a host cell in the recombinant production of a polypeptide ofthe invention. The cell may be transformed with the DNA construct of theinvention, conveniently by integrating the DNA construct in the hostchromosome. This integration is generally considered to be an advantageas the DNA sequence is more likely to be stably maintained in the cell.Integration of the DNA constructs into the host chromosome may beperformed according to conventional methods, e.g. by homologous orheterologous recombination. Alternatively, the cell may be transformedwith an expression vector as described above in connection with thedifferent types of host cells.

The cell of the invention may be a cell of a higher organism such as amammal, an avian, an insect, or a plant cell, but is preferably amicrobial cell, e.g. a bacterial or a fungal (including yeast) cell.

Examples of suitable bacteria are gram positive bacteria such asBacillus subtilis, Bacillus licheniformis, Bacillus lentus, Bacillusbrevis, Bacillus stearothermophilus, Bacillus alkalophilus, Bacillusamyloliguefaciens, Bacillus coagulans, Bacillus circulans, Bacilluslautus, Bacillus megaterium, Bacillus thuringiensis, or Streptomyceslividans or Streptomyces murinus, or gram negative bacteria such as E.coli. The transformation of the bacteria may for instance be effected byprotoplast transformation or by using competent cells in a manner knownper se.

The yeast organism may favourably be selected from a species ofSaccharomyces or Schizosaccharomyces, e.g. Saccharomyces cerevisiae. Thefilamentous fungus may advantageously belong to a species ofAspergillus, e.g. Aspergillus oryzae or Aspergillus niger.Alternatively, a strain of a Fusarium species, e.g. F. oxysporum, can beused as a host cell. Fungal cells may be transformed by a processinvolving protoplast formation and transformation of the protoplastsfollowed by regeneration of the cell wall in a manner known per se. Asuitable procedure for transformation of Aspergillus host cells isdescribed in EP 238 023. A suitable method of transforming Fusariumspecies is described by Malardier et al., 1989.

In a yet further aspect, the present invention relates to a method ofproducing a recombinant protein disulfide isomerase of the invention,which method comprises cultivating a host cell as described above underconditions conducive to the production of the protein disulfideisomerase and recovering the protein disulfide isomerase from the cellsand/or culture medium.

The medium used to cultivate the cells may be any conventional mediumsuitable for growing the host cell in question and obtaining expressionof the protein disulfide isomerase of the invention. Suitable media areavailable from commercial suppliers or may be prepared according topublished recipes (e.g. in catalogues of the American Type CultureCollection).

The resulting protein disulfide isomerase may be recovered from themedium by conventional procedures including separating the cells fromthe medium by centrifugation or filtration, if necessary afterdisruption of the cells, precipitating the proteinaceous components ofthe supernatant or filtrate by means of a salt, e.g. ammonium sulphate,followed by purification by a variety of chromatographic procedures,e.g. ion exchange chromatography, affinity chromatography, or the like.

It is of course also possible to produce the protein disulfideisomerases of the invention by culturing the filamentous fungal naturalhost or parent organism of interest and recovering the protein disulfideisomerase from the culture broth in traditional ways.

The present invention also relates to compositions comprising theprotein disulfide isomerase of the invention.

The compositions may suitably contain 0.01-200 mg of enzyme protein pergram, preferably 0.01-20 mg of enzyme protein per gram, especially0.01-2 mg of enzyme protein per gram, or alternatively 0.02-0.2 mg ofenzyme protein per gram, or 0.01-0.2 mg of enzyme protein per gram.

The compositions of the invention may contain other ingredients known inthe art as e.g. excipients, stabilizers, fillers, detergents, etc.

The compositions of the invention may be formulated in any convenientform, e.g. as a powder, paste, liquid or in granular form. The enzymemay be stabilized in a liquid by inclusion of enzyme stabilizers.Usually, the pH of a solution of the composition of the invention willbe 5-10 and in some instances 7.0-8.5. Other enzymes such as proteases,cellulases, oxidases, peroxidases, amylases or lipases may be includedin the compositions of the invention, either separately or in a combinedadditive.

The compositions of the invention can be used for the treatment ordegradation of scleroproteins, especially hair, skin and wool, dehairingand softening of hides, treatment and cleaning of fabrics, as additivesto detergents, thickening and gelation of food and fodder, strengtheningof gluten in bakery or pastry products, and as pharmaceuticals for thealleviation of eye sufferings.

The present invention is further illustrated in the following exampleswhich should not, in any manner, be considered to limit the scope of thepresent invention.

MATERIALS AND METHODS

Strains

Aspergillus oryzae IFO 4177 available from Institute for Fermentation,Osaka; 17-25 Juso Hammachi 2-Chome Yodogawa-Ku, Osaka, Japan.

Aspergillus niger A524 (ATCC 16882)

Aspergillus niger TSA 1.

E. coli DH5αF'

Determination of PDI activity

The PDI is assayed using the insulin reduction assay described by Jameset al., Cell 67:581-589, 1991.

Plasmids

pUC 19, pMT 1560, pToC 90

EXAMPLES Example 1 Cloning of Aspergillus oryzae and Aspergillus nigerPDI encoding genes

1.1. Design of oligo nucleotides for PCR amplification

PDI from different organisms are highly homologous especially near theactive site residues. In FIG. 1, the following 7 PDI gene products arealigned:

Bovine (Bos taurus) PDI (Yamauchi et al., Biochem. Biophys. Res. Commun.146:1485-1492, 1987), Chicken (Gallus gallus) PDI (Parkkonen et al.,Biochem. J. 256:1005-1011, 1988), Human (Homo sapiens) PDI(Rapilajaniemi et al. EMBO J. 6:643-649, 1987), Mouse (Mus musculus) PDI(Gong, et al., Nucleic Acids Res. 16:1203, 1988), Rabbit (Oryctolaguscuniculus) PDI (Fliegel et al., J. Biol. Chem. 265:15496-15502, 1990),Rat (Rattus norvegicus) PDI (Edman et al., Nature 317:267-270, 1985),Yeast (Saccharomyces cerevisiae) PDI (Tachikawa et al., J. Biochem.110:306-313).

Each subunit contains two active centres (Freedman et al., Cell57:1069-1072, 1989) and the homology in the surroundings of these activecentres are particularly strong. A consensus amino acid sequence for theactive centre closest to the N-terminus was determined from thealignment as -APWCGHCK- (SEQ ID NOS:33-38 respectively, and an oligodeoxyribonucleotide encoding the peptide -WCGHCK- (SEQ ID NOS:24) andextended with an EcoRI site in the 5' end, was synthesized:

5'TGGAATTCTGGTGYGGNCAYTGYAA3' (primer 4762, 25 nucleotides, 32 species,SEQ ID No. 13) (Y=C or T; R=A or G; N=A, T, C, or G).

A consensus amino acid sequence for the active centre closest to theC-terminus was determined: -YAPWCGHCK- (SEQ ID NO:24), and an oligodeoxyribonucleotide encoding the peptide -YAPWCG- (SEQ ID NO:24) inantisense and extended with a BamHI site in the 5' end was synthesized:

5'TGGGATCCRCACCANGGNGCRTA3' (primer 4763, 23 nucleotides, 64 species,SEQ ID NO. 14).

These oligo deoxyribonucleotides (primers 4762 and 4763) were used asprimers in a PCR reaction to amplify PDI-encoding gene fragments fromgenomic DNA from A. oryzae and A. niger.

1.2 Amplification and cloning of fragments of PDI-encoding genes

Genomic DNA was prepared from Aspergillus oryzae IFO 4177 andAspergilIus niger A524 as described by Yelton et al. (Proc. Natl. Acad.Sci. U.S.A. 81:1470-1474, 1984).

PCR reaction mixtures contained Taq DNA polymerase buffer supplied byClontech laboratories Inc. and diluted as described, 250 μM of each ofdATP, dCTP, dGTP, and, dTTP, 100 pmol of each of primers 4762 and 4763,and 0.5 μg of genomic DNA of either A. niger or A. oryzae. The totalreaction volume was 0.1 ml, and it was covered with 0.05 ml paraffinoil.

The following program was run on a Cetus Perkin Elmer thermal cycler:

1. cycle: 94° C. for 2 min., (when the temperature reached 94° C. 2.5 Uof Taq DNA polymerase supplied by Clontech laboratories Inc. was added).

10 cycles: 94° C. for 1 min., 50° C. for 1 min., and 72° C. for 2 min.

30 cycles: 94° C. for 1 min., 55° C. for 1 min., and 72° C. for 2 min.

1 cycle: 72° C. for 5 min.

The reaction mixtures were loaded on an agarose gel, and both the A.oryzae and the A. niger DNA produced fragments of approximately 1.1 kb.

The fragments were digested with EcoRI and BamHI and ligated to pUC19(Yanisch-Perron et al., Gene 33:103-119, 1985). The ligation mixture wastransformed into E. coli DH5αF' (Woodcock et al., Nucleic Acids Res.(1989) 17:3469-3478). Recombinant plasmids were subjected to sequenceanalysis using the Sequenase™ kit (United States Biochemical) and a M13universal primer following the manufacturers instructions. The analysisconfirmed that both in the case of A. oryzae and in that of A. nigersequences homologous to other PDI genes were amplified and cloned.

1.3 Genome cloning of the A. oryzae PDI-encoding gene

Genomic DNA from A. oryzae was digested with the following restrictionenzymes supplied by New England Biolabs Inc.: HindIII, BamHI,BamHI+HindIII, EcoRI, EcoRI+HindIII, SalI, SalI+HindIII, BglII,BglII+HindIII, PstI and PstI+HindIII. After digestion, the reactionmixtures were run on a 1% agarose gel and then blotted onto an ImmobilonN™ membrane (Millipore Corporation) following the manufacturersinstructions. The membrane was probed with the cloned A. oryzae PCRproduct isolated as a BamHI-EcoRI fragment and radio labelled with ³² P.After stringent washes the membrane was subjected to autoradiography.

Genomic DNA from A. niger was digested with the following restrictionenzymes: BglII, BamHI, BamHI+BglII, EcoRI, EcoRI+BglII, SalI,SalI+BglII, HindIII, HindIII+BglII, PstI and PstI+BglII. The Southernblot was made as described with A. oryzae, only the A. niger PCR productwas used as probe.

1.4 Construction of genomic A. oryzae library

Southern analysis indicated that the A. oryzae PDI gene was located on a6.8 kb BglII fragment. Genomic A. oryzae DNA was digested with BglII andfragments ranging from 5 kb to 8.5 kb were isolated from an agarose gel.Subcloning thereof and Southern analysis indicated that the A. oryzaePDI gene was located on a 2.3 kb BamHI, HindIII fragment. Genomic A.oryzae DNA was digested with BamHI and HindIII and fragments rangingfrom 1.9-3 kb were isolated from an agarose gel. This mixture offragments was ligated to pUC19 digested with BamHI and HindIII. Theligation mixture was used to transform E. coli DH5αF'. The transformedE. coli cells were spread onto 10 agar plates using ampicillinselection.

1.5 Screening of the A. oryzae genomic library

The libraries were screened using the filter colony hybridization methoddescribed by Gergen et al. (Nucleic Acids Res. 7:2115-2136, 1979). Theprobe that was used for the Southern blot was also used for the colonyhybridization. Positive clones were isolated and confirmed by sequenceanalysis using sequencing primers designed from the sequences of the PDIfragments. One of the plasmids containing the desired fragment wastermed pCaHj 425.

1.6 Sequence of the gene

The gene was sequenced using the Tag DyeDeoxy™ Terminator cyclesequencing kit supplied by Applied Biosystems following themanufacturer's instructions. The sequence reactions were run on anApplied Biosystems 373A DNA sequencer and the data were evaluated usingthe Macintosh computer program SegEd version 1.0 supplied by AppliedBiosystems.

The sequence of the A. oryzae gene is shown in the appended SEQ ID No.1.

The amino acid composition of the purified PDI obtained as described inExample 2 was in accordance with the composition deduced from theDNA-sequence shown in SEQ ID No. 1. From homology to other PDI genes andconsensus splicing sequences a cDNA sequence as shown in SEQ ID no. 2was suggested. The derived protein sequence is as shown in SEQ ID No. 3.

Example 2 Expression of truncated forms of the A. oryzae PDI gene

2.1 Construction of expression plasmids

The PDI gene of A. oryzae was truncated at various positions byintroduction stop codons. This was done by PCR amplification of the PDIgene using a 5' PCR primer harbouring a BamHI site at its 5' end and 8different 3' primers corresponding to 8 different truncations eachharbouring a HindIII site. The sequence of the 5' primer was: ##STR1##

The sequences of the eight 3' primers were: ##STR2## Primer 5215directed an extension of the PDI gene amino acid 1-115 with the sequenceLeu-Ile-Arg-Glu-Leu-Leu-Gln-Glu-Leu-Val-Asn-Lys-His-Leu (SEQ ID NO:11);followed by a stop codon.

Primer 5397 introduced a stop codon after amino acid 131.

Primer 5895 introduced a stop codon after amino acid 141.

Primer 5399 introduced a stop codon after amino acid 143.

Primer 5894 introduced a stop codon after amino acid 163.

Primer 5893 introduced a stop codon after amino acid 174.

Primer 6314 introduced a stop codon after amino acid 281.

Primer 5204 introduced the mutation E511A (meaning substituting and astop codon after amino acid 511.

The expression plasmids were constructed by PCR amplification usingprimer 5205 in combination with either 5215, 5397, 5895, 5399, 5894,5893, 6314 or 5204 and pCaHj 425 as template using standard PCRconditions. The generated PCR fragments were digested with BamHI andHindIII and inserted into pMT 1560 (described in e.g. PCT/DK94/00138)digested with the same enzymes (See FIG. 3). The constructed plasmidswere named pCaHj 432 (from primer 5215), pCaHj 433 (from primer 5397),PCaHj 441 (from primer 5895), pCaHj 434 (from primer 5399), pCaHj 440(from primer 5894), pCaHj 439 (from primer 5893), pCaHj 445 (from primer6314) and pCaHj 431 (from primer 5204).

2.2 Transformation of A. oryzae IFO 4177.

Each of the plasmids pCaHj 432, pCaHj 433, PCaHj 441, pCaHj 434, pCaHj440, pCaHj 439, pCaHj 445 and pCaHj 431 were transformed into A. oryzaeIFO 4177 by cotransformation with the amdS selection plasmid pToC 90(described in WO91/17243) following the procedure described in thepublished EP patent application No. 238 023.

A number of transformants of each plasmid were evaluated.

2.3 Transformation of A. niger TSA 1.

Each of the plasmids PCaHj 441, pCaHj 434, pCaHj 440 and pCaHj 439 weretransformed into A. niger TSA 1 by the same procedure as with A. oryzae.

A number of transformants of each plasmid were evaluated.

Example 3 Fermentation purification and characterization of theAspergillus oryzae PDI truncations

3.1 A. oryzae IFO 4177 transformants.

Crude truncated PDI preparation was isolated from supernatants obtainedby fermentation of the A. oryzae or A. niger pCaHj 432, pCaHj 433, PCaHj441, pCaHj 434, pCaHj 440, pCaHj 439, pCaHj 445 or pCaHj 431transformants in shake flasks containing YPM medium (1 liter: 5 g DifcoYeast extract, 10 g Difco peptone, 20 g maltose). The supernatant wasrecovered by filtration. The PDI truncation gene products were partiallypurified using a 1 ml HiTrap Q™ anion exchanger from Pharmacia LKBBiotechnology AB Uppsala, Sweden following the manufacturersinstructions. Fractions were collected and analyzed by measuring thedisulphide isomerase activity and by SDS PAGE.

The pCaHj 434 transformants secreted a protein of approx 14 kD (SDSPAGE) not present in supernatants of the untransformed strain.Enrichment of this protein by ion exchange was followed by increaseddisulphide isomerase activity. The approx. 14 kD band was blotted froman SDS Page gel and subjected to N-terminal amino acid sequencedetermination using an Applied Biosystems 473A protein sequencer. Asequence of 7 amino acids could unambiguously be determined as:Thr-Ala-Glu-Ala-Pro-Ser-Asp (SEQ ID NO:25). This sequence corresponds toresidue 24-30 of the A. oryzae protein sequence. The size of thetruncation expected from the amino acid sequence is thus 13.2 kD. So itcan be concluded that the pCaHj 434 gene product is secreted to thesupernatant, that it has the expected size and that it is catalyticactive.

The pCaHj 441 transformants secreted a protein of the same size as thepCaHj 434 transformants. Also for this truncation enrichment of theprotein was followed by increased disulphide isomerase activitydemonstrating that the pCaHj 441 gene product is a catalytic activesecreted protein.

The pCaHj 440 transformants secreted a protein of approx 16 kD notpresent in the untransformed strain. The expected size is 15.7 kDassuming the same N-terminal sequence as the pCaHj 434 product.Enrichment of the protein by ion exchange was followed by increaseddisulphide isomerase activity demonstrating that also the pCaHj 440 geneproduct is a catalytic active secreted protein.

The pCaHj 445 transformants secreted a protein of approx 30 kD notpresent in the untransformed strain. The expected size is 28.6 kDassuming the same N-terminal sequence as the pCaHj 434 product.Enrichment of the protein by ion exchange was followed by increaseddisulphide isomerase activity demonstrating that the pCaHj 440 geneproduct is a catalytic active secreted protein.

3.2 A. niger TSA 1 transformants.

Transformants of PCaHj 441, pCaHj 434, pCaHj 440 and pCaHj 439 wereevaluated in the same way as the corresponding A. oryzae transformantswith the exception that the N-terminal amino acid sequence was notdetermined for any of the proteins secreted by A. niger.

In all other aspects the same results were obtained with the A. nigertransformants as with the A. oryzae transformants. However thefermentation yield of the truncations were generally lower in A. nigerthan in A. oryzae.

                                      TABLE 1    __________________________________________________________________________    (SEQ ID NOS: 26-32)    __________________________________________________________________________     ##STR3##     ##STR4##     ##STR5##     ##STR6##     ##STR7##     ##STR8##     ##STR9##     ##STR10##     ##STR11##     ##STR12##     ##STR13##    __________________________________________________________________________

                                      TABLE 2    __________________________________________________________________________    (SEQ ID NOS 33-38)    __________________________________________________________________________     ##STR14##     ##STR15##     ##STR16##     ##STR17##     ##STR18##     ##STR19##     ##STR20##     ##STR21##     ##STR22##     ##STR23##     ##STR24##     ##STR25##     ##STR26##     ##STR27##    __________________________________________________________________________

REFERENCES CITED IN THE SPECIFICATION

Sambrook, J., Fritsch, E. F. & Maniatis, T. 1989. Molecular Cloning: ALaboratory Manual. 2. edition. Cold Spring Harbor Lab., Cold SpringHarbor, N.Y.

U.S. Pat. No. 4,683,202

Hudson et al, 1989, Practical Immunology, Third edition, BlackwellScientific Publications.

Beaucage and Caruthers, 1981, Tetrahedron Letters 22, 1981, pp.1859-1869

Matthes et al., 1984, The EMBO J. 3, 1984, pp. 801-805

R. K. Saiki et al., 1988, Science 239, pp. 487-491

WO 91/17243

EP 238 023

Malardier et al. Gene 78 (1989), pp. 147-156

Woodcock et al., Nucleic Acids Res. (1989) 17:3469-3478

    __________________________________________________________________________    SEQUENCE LISTING    (1) GENERAL INFORMATION:    (iii) NUMBER OF SEQUENCES: 38    (2) INFORMATION FOR SEQ ID NO: 1:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 1953 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: DNA (genomic)    (iii) HYPOTHETICAL: NO    (iii) ANTI-SENSE: NO    (vi) ORIGINAL SOURCE:    (A) ORGANISM: Aspergillus oryzae    (B) STRAIN: IFO 4177    (ix) FEATURE:    (A) NAME/KEY: CDS    (B) LOCATION: join(71..445, 503..880, 962..1402,    1479..1829)    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 1:    CCCTGCTGTCCCCATAGACAGTACACACGTCATCCTTTGATATTGTCACACTTGACAAAT60    TCCCGACACCATGCGGACTTTCGCACCTTGGATCTTGAGCCTTCTAGGG109    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGly    1510    GCTTCTGCTGTAGCTTCTGCTGCCGATGCGACTGCCGAAGCTCCCTCC157    AlaSerAlaValAlaSerAlaAlaAspAlaThrAlaGluAlaProSer    152025    GATGTGGTCTCGCTCACCGGGGACACATTCGAAACTTTCGTCAAGGAG205    AspValValSerLeuThrGlyAspThrPheGluThrPheValLysGlu    30354045    CATGACCTAGTTTTGGCCGAGTTTTTTGCTCCCTGGTGTGGCCATTGC253    HisAspLeuValLeuAlaGluPhePheAlaProTrpCysGlyHisCys    505560    AAGGCTCTCGCTCCGAAATACGAGCAGGCCGCCACTGAGTTAAAGGAA301    LysAlaLeuAlaProLysTyrGluGlnAlaAlaThrGluLeuLysGlu    657075    AAGAACATTCCGCTGGTCAAGGTTGATTGCACCGAGGAAGAGGCTCTT349    LysAsnIleProLeuValLysValAspCysThrGluGluGluAlaLeu    808590    TGTAGGGACCAAGGTGTTGAAGGTTACCCCACGCTGAAGATTTTCCGT397    CysArgAspGlnGlyValGluGlyTyrProThrLeuLysIlePheArg    95100105    GGCCTTGACGCTGTTAAGCCTTATCAGGGAGCTCGTCAGACCGAGGCG445    GlyLeuAspAlaValLysProTyrGlnGlyAlaArgGlnThrGluAla    110115120125    GTAAGTGTCACCTGTTTGTTAGCCTTGCTCAAATAATATTGACCGCTAGTATCATAG502    ATTGTTTCATACATGGTCAAGCAGTCACTACCTGCTGTGTCCCCTGTC550    IleValSerTyrMetValLysGlnSerLeuProAlaValSerProVal    130135140    ACCCCAGAAAACCTCGAAGAGATCAAGACTATGGACAAGATTGTCGTT598    ThrProGluAsnLeuGluGluIleLysThrMetAspLysIleValVal    145150155    ATTGGTTATATCGCGTCTGACGACCAGACTGCCAATGATATATTCACC646    IleGlyTyrIleAlaSerAspAspGlnThrAlaAsnAspIlePheThr    160165170    ACTTTTGCCGAGTCACAGAGAGACAACTACCTCTTCGCCGCCACAAGT694    ThrPheAlaGluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSer    175180185    GATGCATCGATCGCTAAGGCAGAAGGTGTTAAGCAACCTTCGATTGTT742    AspAlaSerIleAlaLysAlaGluGlyValLysGlnProSerIleVal    190195200205    CTCTATAAAGACTTCGATGAAAAGAAAGCTACTTATGATGGAGAGATT790    LeuTyrLysAspPheAspGluLysLysAlaThrTyrAspGlyGluIle    210215220    GAACAGGATGCCCTCCTCAGTTGGGTCAAGACTGCCAGTACCCCCTTG838    GluGlnAspAlaLeuLeuSerTrpValLysThrAlaSerThrProLeu    225230235    GTGGGCGAGCTGGGCCCAGAGACTTACTCCGGATATATAACG880    ValGlyGluLeuGlyProGluThrTyrSerGlyTyrIleThr    240245250    GTATGTCACAAGACACAATCTCAATATCGCTTCACAACGTTTAGTAAATAATCATGAGTT940    TCTGACATGGGTTTGGTTAAGGCTGGCATTCCACTGGCGTACATTTTCGCC991    AlaGlyIleProLeuAlaTyrIlePheAla    255260    GAAACCAAAGAAGAGCGTGAGCAGTTCACCGAGGAGTTCAAGTTCATC1039    GluThrLysGluGluArgGluGlnPheThrGluGluPheLysPheIle    265270275    GCCGAGAAACACAAGGGTTCCATCAATATTGTCACCATTGACGCCAAG1087    AlaGluLysHisLysGlySerIleAsnIleValThrIleAspAlaLys    280285290    TTGTACGGCGCTCATGCAGGCAATCTCAACCTTGACCCCTCCAAGTTC1135    LeuTyrGlyAlaHisAlaGlyAsnLeuAsnLeuAspProSerLysPhe    295300305    CCTGCATTCGCTATTCAAGACCCTGAAAAGAACGCCAAGTATCCTTAT1183    ProAlaPheAlaIleGlnAspProGluLysAsnAlaLysTyrProTyr    310315320325    GACCAGTCGAAGGAAGTCAAGGCCAAGGATATCGGTAAATTCATCCAA1231    AspGlnSerLysGluValLysAlaLysAspIleGlyLysPheIleGln    330335340    GACGTTCTTGATGATAAAGTAGAGCCAAGCATTAAGTCTGAGGCTATT1279    AspValLeuAspAspLysValGluProSerIleLysSerGluAlaIle    345350355    CCTGAGACTCAGGAAGGTCCTGTTACTGTTGTTGTCGCGCATTCCTAT1327    ProGluThrGlnGluGlyProValThrValValValAlaHisSerTyr    360365370    AAGGATCTCGTCCTTGACAACGAGAAGGACGTCCTTCTCGAATTTTAT1375    LysAspLeuValLeuAspAsnGluLysAspValLeuLeuGluPheTyr    375380385    GCGCCATGGTGCGGACACTGCAAGGCCGTAAGTTTTCCCCCTCTTTC1422    AlaProTrpCysGlyHisCysLysAla    390395    TCTACAACGAATTATATCCACTCTCGCTTGCGAATACCTAATTAAACCTTGAATAG1478    CTTGCCCCGAAGTACGAGGAACTTGCAAGCCTTTACAAGGATATTCCT1526    LeuAlaProLysTyrGluGluLeuAlaSerLeuTyrLysAspIlePro    400405410    GAAGTTACCATCGCCAAAATTGACGCAACGGCCAACGATGTCCCCGAC1574    GluValThrIleAlaLysIleAspAlaThrAlaAsnAspValProAsp    415420425430    TCCATTACAGGATTTCCTACTATTAAGCTCTTCGCTGCCGGCGCCAAG1622    SerIleThrGlyPheProThrIleLysLeuPheAlaAlaGlyAlaLys    435440445    GACTCCCCAGTTGAATATGAAGGCTCTCGCACGGTGGAGGACCTCGCC1670    AspSerProValGluTyrGluGlySerArgThrValGluAspLeuAla    450455460    AACTTCGTCAAGGAGAATGGCAAGCACAAGGTCGATGCTCTTGAAGTT1718    AsnPheValLysGluAsnGlyLysHisLysValAspAlaLeuGluVal    465470475    GATCCGAAGAAAGAACAGGAGAGTGGCGATGCCACCGAGACTCGGGCC1766    AspProLysLysGluGlnGluSerGlyAspAlaThrGluThrArgAla    480485490    GCCTCTGACGAGACCGAAACTCCTGCTGCTACTAGCGATGACAAGTCT1814    AlaSerAspGluThrGluThrProAlaAlaThrSerAspAspLysSer    495500505510    GAGCATGATGAATTGTAAATTTCATTTGGCCTGATAGTTTGATCCATATTTATGT1869    GluHisAspGluLeu    515    GAATTCTTGTATTCTACCAGCAGTTTGAGCAATCGCAGCTACTTCCGGCTTAGGAAACTG1929    TTGTTCTATCCTAGTGGGAAGCTT1953    (2) INFORMATION FOR SEQ ID NO: 2:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 1547 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: cDNA    (iii) HYPOTHETICAL: NO    (iii) ANTI-SENSE: NO    (vi) ORIGINAL SOURCE:    (A) ORGANISM: Aspergillus oryzae    (B) STRAIN: IFO 4177    (ix) FEATURE:    (A) NAME/KEY: CDS    (B) LOCATION: 1..1547    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 2:    ATGCGGACTTTCGCACCTTGGATCTTGAGCCTTCTAGGGGCTTCTGCT48    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    GTAGCTTCTGCTGCCGATGCGACTGCCGAAGCTCCCTCCGATGTGGTC96    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    TCGCTCACCGGGGACACATTCGAAACTTTCGTCAAGGAGCATGACCTA144    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    GTTTTGGCCGAGTTTTTTGCTCCCTGGTGTGGCCATTGCAAGGCTCTC192    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    GCTCCGAAATACGAGCAGGCCGCCACTGAGTTAAAGGAAAAGAACATT240    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    CCGCTGGTCAAGGTTGATTGCACCGAGGAAGAGGCTCTTTGTAGGGAC288    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    CAAGGTGTTGAAGGTTACCCCACGCTGAAGATTTTCCGTGGCCTTGAC336    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    GCTGTTAAGCCTTATCAGGGAGCTCGTCAGACCGAGGCGATTGTTTCA384    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TACATGGTCAAGCAGTCACTACCTGCTGTGTCCCCTGTCACCCCAGAA432    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AACCTCGAAGAGATCAAGACTATGGACAAGATTGTCGTTATTGGTTAT480    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    ATCGCGTCTGACGACCAGACTGCCAATGATATATTCACCACTTTTGCC528    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThrPheAla    165170175    GAGTCACAGAGAGACAACTACCTCTTCGCCGCCACAAGTGATGCATCG576    GluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSerAspAlaSer    180185190    ATCGCTAAGGCAGAAGGTGTTAAGCAACCTTCGATTGTTCTCTATAAA624    IleAlaLysAlaGluGlyValLysGlnProSerIleValLeuTyrLys    195200205    GACTTCGATGAAAAGAAAGCTACTTATGATGGAGAGATTGAACAGGAT672    AspPheAspGluLysLysAlaThrTyrAspGlyGluIleGluGlnAsp    210215220    GCCCTCCTCAGTTGGGTCAAGACTGCCAGTACCCCCTTGGTGGGCGAG720    AlaLeuLeuSerTrpValLysThrAlaSerThrProLeuValGlyGlu    225230235240    CTGGGCCCAGAGACTTACTCCGGATATATAACGGCTGGCATTCCACTG768    LeuGlyProGluThrTyrSerGlyTyrIleThrAlaGlyIleProLeu    245250255    GCGTACATTTTCGCCGAAACCAAAGAAGAGCGTGAGCAGTTCACCGAG816    AlaTyrIlePheAlaGluThrLysGluGluArgGluGlnPheThrGlu    260265270    GAGTTCAAGTTCATCGCCGAGAAACACAAGGGTTCCATCAATATTGTC864    GluPheLysPheIleAlaGluLysHisLysGlySerIleAsnIleVal    275280285    ACCATTGACGCCAAGTTGTACGGCGCTCATGCAGGCAATCTCAACCTT912    ThrIleAspAlaLysLeuTyrGlyAlaHisAlaGlyAsnLeuAsnLeu    290295300    GACCCCTCCAAGTTCCCTGCATTCGCTATTCAAGACCCTGAAAAGAAC960    AspProSerLysPheProAlaPheAlaIleGlnAspProGluLysAsn    305310315320    GCCAAGTATCCTTATGACCAGTCGAAGGAAGTCAAGGCCAAGGATATC1008    AlaLysTyrProTyrAspGlnSerLysGluValLysAlaLysAspIle    325330335    GGTAAATTCATCCAAGACGTTCTTGATGATAAAGTAGAGCCAAGCATT1056    GlyLysPheIleGlnAspValLeuAspAspLysValGluProSerIle    340345350    AAGTCTGAGGCTATTCCTGAGACTCAGGAAGGTCCTGTTACTGTTGTT1104    LysSerGluAlaIleProGluThrGlnGluGlyProValThrValVal    355360365    GTCGCGCATTCCTATAAGGATCTCGTCCTTGACAACGAGAAGGACGTC1152    ValAlaHisSerTyrLysAspLeuValLeuAspAsnGluLysAspVal    370375380    CTTCTCGAATTTTATGCGCCATGGTGCGGACACTGCAAGGCCCTTGCC1200    LeuLeuGluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAla    385390395400    CCGAAGTACGAGGAACTTGCAAGCCTTTACAAGGATATTCCTGAAGTT1248    ProLysTyrGluGluLeuAlaSerLeuTyrLysAspIleProGluVal    405410415    ACCATCGCCAAAATTGACGCAACGGCCAACGATGTCCCCGACTCCATT1296    ThrIleAlaLysIleAspAlaThrAlaAsnAspValProAspSerIle    420425430    ACAGGATTTCCTACTATTAAGCTCTTCGCTGCCGGCGCCAAGGACTCC1344    ThrGlyPheProThrIleLysLeuPheAlaAlaGlyAlaLysAspSer    435440445    CCAGTTGAATATGAAGGCTCTCGCACGGTGGAGGACCTCGCCAACTTC1392    ProValGluTyrGluGlySerArgThrValGluAspLeuAlaAsnPhe    450455460    GTCAAGGAGAATGGCAAGCACAAGGTCGATGCTCTTGAAGTTGATCCG1440    ValLysGluAsnGlyLysHisLysValAspAlaLeuGluValAspPro    465470475480    AAGAAAGAACAGGAGAGTGGCGATGCCACCGAGACTCGGGCCGCCTCT1488    LysLysGluGlnGluSerGlyAspAlaThrGluThrArgAlaAlaSer    485490495    GACGAGACCGAAACTCCTGCTGCTACTAGCGATGACAAGTCTGAGCAT1536    AspGluThrGluThrProAlaAlaThrSerAspAspLysSerGluHis    500505510    GATGAATTGTA1547    AspGluLeu    515    (2) INFORMATION FOR SEQ ID NO: 3:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 515 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 3:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThrPheAla    165170175    GluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSerAspAlaSer    180185190    IleAlaLysAlaGluGlyValLysGlnProSerIleValLeuTyrLys    195200205    AspPheAspGluLysLysAlaThrTyrAspGlyGluIleGluGlnAsp    210215220    AlaLeuLeuSerTrpValLysThrAlaSerThrProLeuValGlyGlu    225230235240    LeuGlyProGluThrTyrSerGlyTyrIleThrAlaGlyIleProLeu    245250255    AlaTyrIlePheAlaGluThrLysGluGluArgGluGlnPheThrGlu    260265270    GluPheLysPheIleAlaGluLysHisLysGlySerIleAsnIleVal    275280285    ThrIleAspAlaLysLeuTyrGlyAlaHisAlaGlyAsnLeuAsnLeu    290295300    AspProSerLysPheProAlaPheAlaIleGlnAspProGluLysAsn    305310315320    AlaLysTyrProTyrAspGlnSerLysGluValLysAlaLysAspIle    325330335    GlyLysPheIleGlnAspValLeuAspAspLysValGluProSerIle    340345350    LysSerGluAlaIleProGluThrGlnGluGlyProValThrValVal    355360365    ValAlaHisSerTyrLysAspLeuValLeuAspAsnGluLysAspVal    370375380    LeuLeuGluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAla    385390395400    ProLysTyrGluGluLeuAlaSerLeuTyrLysAspIleProGluVal    405410415    ThrIleAlaLysIleAspAlaThrAlaAsnAspValProAspSerIle    420425430    ThrGlyPheProThrIleLysLeuPheAlaAlaGlyAlaLysAspSer    435440445    ProValGluTyrGluGlySerArgThrValGluAspLeuAlaAsnPhe    450455460    ValLysGluAsnGlyLysHisLysValAspAlaLeuGluValAspPro    465470475480    LysLysGluGlnGluSerGlyAspAlaThrGluThrArgAlaAlaSer    485490495    AspGluThrGluThrProAlaAlaThrSerAspAspLysSerGluHis    500505510    AspGluLeu    515    (2) INFORMATION FOR SEQ ID NO: 4:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 511 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 4:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThrPheAla    165170175    GluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSerAspAlaSer    180185190    IleAlaLysAlaGluGlyValLysGlnProSerIleValLeuTyrLys    195200205    AspPheAspGluLysLysAlaThrTyrAspGlyGluIleGluGlnAsp    210215220    AlaLeuLeuSerTrpValLysThrAlaSerThrProLeuValGlyGlu    225230235240    LeuGlyProGluThrTyrSerGlyTyrIleThrAlaGlyIleProLeu    245250255    AlaTyrIlePheAlaGluThrLysGluGluArgGluGlnPheThrGlu    260265270    GluPheLysPheIleAlaGluLysHisLysGlySerIleAsnIleVal    275280285    ThrIleAspAlaLysLeuTyrGlyAlaHisAlaGlyAsnLeuAsnLeu    290295300    AspProSerLysPheProAlaPheAlaIleGlnAspProGluLysAsn    305310315320    AlaLysTyrProTyrAspGlnSerLysGluValLysAlaLysAspIle    325330335    GlyLysPheIleGlnAspValLeuAspAspLysValGluProSerIle    340345350    LysSerGluAlaIleProGluThrGlnGluGlyProValThrValVal    355360365    ValAlaHisSerTyrLysAspLeuValLeuAspAsnGluLysAspVal    370375380    LeuLeuGluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAla    385390395400    ProLysTyrGluGluLeuAlaSerLeuTyrLysAspIleProGluVal    405410415    ThrIleAlaLysIleAspAlaThrAlaAsnAspValProAspSerIle    420425430    ThrGlyPheProThrIleLysLeuPheAlaAlaGlyAlaLysAspSer    435440445    ProValGluTyrGluGlySerArgThrValGluAspLeuAlaAsnPhe    450455460    ValLysGluAsnGlyLysHisLysValAspAlaLeuGluValAspPro    465470475480    LysLysGluGlnGluSerGlyAspAlaThrGluThrArgAlaAlaSer    485490495    AspGluThrGluThrProAlaAlaThrSerAspAspLysSerAla    500505510    (2) INFORMATION FOR SEQ ID NO: 5:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 281 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 5:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThrPheAla    165170175    GluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSerAspAlaSer    180185190    IleAlaLysAlaGluGlyValLysGlnProSerIleValLeuTyrLys    195200205    AspPheAspGluLysLysAlaThrTyrAspGlyGluIleGluGlnAsp    210215220    AlaLeuLeuSerTrpValLysThrAlaSerThrProLeuValGlyGlu    225230235240    LeuGlyProGluThrTyrSerGlyTyrIleThrAlaGlyIleProLeu    245250255    AlaTyrIlePheAlaGluThrLysGluGluArgGluGlnPheThrGlu    260265270    GluPheLysPheIleAlaGluLysHis    275280    (2) INFORMATION FOR SEQ ID NO: 6:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 174 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 6:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThr    165170    (2) INFORMATION FOR SEQ ID NO: 7:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 163 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 7:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSer    (2) INFORMATION FOR SEQ ID NO: 8:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 143 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 8:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrPro    130135140    (2) INFORMATION FOR SEQ ID NO: 9:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 141 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 9:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProVal    130135140    (2) INFORMATION FOR SEQ ID NO: 10:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 131 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 10:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetVal    130    (2) INFORMATION FOR SEQ ID NO: 11:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 129 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 11:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysLeuIleArgGluLeuLeuGlnGluLeuValAsnLysHis    115120125    Leu    (2) INFORMATION FOR SEQ ID NO: 12:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 200 amino acids    (B) TYPE: amino acid    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: protein    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 12:    AlaGluAlaProSerAspValValSerLeuThrGlyAspThrPheGlu    151015    ThrPheValLysGluHisAspLeuValLeuAlaGluPhePheAlaPro    202530    TrpCysGlyHisCysLysAlaLeuAlaProLysTyrGluGlnAlaAla    354045    ThrGluLeuLysGluLysAsnIleProLeuValLysValAspCysThr    505560    GluGluGluAlaLeuCysArgAspGlnGlyValGluGlyTyrProThr    65707580    LeuLysIlePheArgGlyLeuAspAlaValLysProTyrGlnGlyAla    859095    ArgGlnThrGluAlaIleValSerTyrMetValLysGlnSerLeuPro    100105110    AlaValSerProValThrProGluAsnLeuGluGluIleLysThrMet    115120125    AspLysIleValValIleGlyTyrIleAlaSerAspAspGlnThrAla    130135140    AsnAspIlePheThrThrPheAlaGluSerGlnArgAspAsnTyrLeu    145150155160    PheAlaAlaThrSerAspAlaSerIleAlaLysAlaGluGlyValLys    165170175    GlnProSerIleValLeuTyrLysAspPheAspGluLysLysAlaThr    180185190    TyrAspGlyGluIleGluGlnAsp    195200    (2) INFORMATION FOR SEQ ID NO: 13:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 25 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 4762    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 13:    TGGAATTCTGGTGYGGNCAYTGYAA25    (2) INFORMATION FOR SEQ ID NO: 14:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 23 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 4763    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 14:    TGGGATCCRCACCANGGNGCRTA23    (2) INFORMATION FOR SEQ ID NO: 15:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 29 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5205    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 15:    TTCGGATCCACCATGCGGACTTTCGCACC29    (2) INFORMATION FOR SEQ ID NO: 16:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 55 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5215    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 16:    CCAAGCTTTAGAGATGCTTGTTGACAAGCTCCTGGAGGAGCTCCCTGATAAGCTT55    (2) INFORMATION FOR SEQ ID NO: 17:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 45 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5397    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 17:    CCAAGCTTTAGACCATGTATGACACAATCGCCTCGGTCTGACGAG45    (2) INFORMATION FOR SEQ ID NO: 18:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 31 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5895    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 18:    CCAAGCTTTAGACAGGGGACACAGCAGGTAG31    (2) INFORMATION FOR SEQ ID NO: 19:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 27 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5399    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 19:    CCAAGCTTTATGGGGTGACAGGGGACA27    (2) INFORMATION FOR SEQ ID NO: 20:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 31 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5894    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 20:    CCAAGCTTTAAGACGCGATATAACCAATAAC31    (2) INFORMATION FOR SEQ ID NO: 21:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 31 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5893    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 21:    CCAAGCTTTAAGTGGTGAATATATCATTGGC31    (2) INFORMATION FOR SEQ ID NO: 22:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 30 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 6314    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 22:    CCAAGCTTAGTGTTTCTCGGCGATGAACTT30    (2) INFORMATION FOR SEQ ID NO: 23:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 31 base pairs    (B) TYPE: nucleic acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: primer 5204    (iii) HYPOTHETICAL: YES    (iii) ANTI-SENSE: NO    (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 23:    CCAAGCTTTACGCAGACTTGTCATCGCTAGT31    (2) INFORMATION FOR SEQ ID NO:24:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 8 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24:    AlaProTrpCysGlyHisCysLys    15    (2) INFORMATION FOR SEQ ID NO:25:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 7 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:    ThrAlaGluAlaProSerAsp    15    (2) INFORMATION FOR SEQ ID NO:26:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 3052 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:26:    MetLeuSerArgAlaLeuLeuCysLeuAlaLeuAlaTrpAlaAlaArg    151015    ValGlyAlaAspAlaLeuGluGluGluAspAsnValLeuValLeuLys    202530    LysSerAsnPheAlaGluAlaLeuAlaAlaHisMetLeuArgArgAla    354045    LeuLeuCysLeuAlaLeuThrAlaLeuPheArgAlaGlyAlaGlyAla    505560    ProAspGluGluAspHisValLeuValLeuHisLysGlyAsnPheAsp    65707580    GluAlaLeuAlaAlaHisMetLeuArgArgAlaLeuLeuCysLeuAla    859095    ValAlaAlaLeuValArgAlaAspAlaProGluGluGluAspHisVal    100105110    LeuValLeuArgLysSerAsnPheAlaGluAlaLeuAlaAlaHisMet    115120125    LeuArgArgAlaValLeuCysLeuAlaLeuAlaValThrAlaGlyTrp    130135140    AlaTrpAlaAlaGluGluGluAspAsnValLeuValLeuLysSerSer    145150155160    AsnPheAlaGluGluLeuAlaAlaHisGluProLeuGluGluGluAsp    165170175    GlyValLeuValLeuArgAlaAlaAsnPheGluGlnAlaLeuAlaAla    180185190    HisMetLysPheSerAlaGlyAlaValLeuSerTrpSerSerLeuLeu    195200205    LeuAlaSerSerValPheAlaGlnGlnGluAlaValAlaProGluAsp    210215220    SerAlaValValLysLeuAlaThrAspSerPheAsnGluTyrIleGln    225230235240    SerHisAsnTyrLeuLeuValGluPheTyrAlaProTrpCysGlyHis    245250255    CysLysAlaLeuAlaProGluTyrAlaLysAlaAlaAlaLysLeuLys    260265270    AlaGluGlySerGluIleArgLeuAlaLysValAspAlaThrGluGlu    275280285    SerAspLeuAlaLysTyrLeuLeuValGluPheTyrAlaProTrpCys    290295300    GlyHisCysLysAlaLeuAlaProGluTyrAlaLysAlaAlaGlyLys    305310315320    LeuLysAlaGluGlySerGluIleArgLeuAlaLysValAspAlaThr    325330335    GluGluSerAspLeuAlaLysTyrLeuLeuValGluPheTyrAlaPro    340345350    TrpCysGlyHisCysLysAlaLeuAlaProGluTyrAlaLysAlaAla    355360365    GlyLysLeuLysAlaGluGlySerGluIleArgLeuAlaLysValAsp    370375380    AlaThrGluGluSerAspLeuAlaLysTyrLeuLeuValGluPheTyr    385390395400    AlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyrAlaLys    405410415    AlaAlaGlyLysLeuLysAlaGluGlySerAspIleArgLeuAlaLys    420425430    ValAspAlaThrGluGluSerAspLeuAlaArgHisLeuLeuValGlu    435440445    PheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyr    450455460    AlaLysAlaAlaAlaGlnLeuLysAlaGluGlySerGluIleArgLeu    465470475480    AlaLysValAspAlaThrGluGluAlaGluLeuAlaAspLeuValLeu    485490495    AlaGluPhePheAlaProTrpCysGlyHisCysLysAsnMetAlaPro    500505510    GluTyrValLysAlaAlaGluThrLeuValGluLysAsnIleThrLeu    515520525    AlaGlnIleAspCysThrGluAsnGlnAspLeuCysGlnGlnTyrGly    530535540    ValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThrAla    545550555560    SerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleValAsn    565570575    TrpLeuLysLysArgThrGlyProAlaAlaThrThrLeuSerGlnGln    580585590    TyrGlyValArgGlyTyrProThrIleLysPhePheLysAsnGlyAsp    595600605    ThrAlaSerProLysGluTyrThrAlaGlyArgGluAlaAspAspIle    610615620    ValAsnTrpLeuLysLysArgThrGlyProAlaAlaSerThrLeuSer    625630635640    GlnGlnTyrGlyValArgGlyTyrProThrIleLysPhePheArgAsn    645650655    GlyAspThrAlaSerProLysGluTyrThrAlaGlyArgGluAlaAsp    660665670    AspIleValAsnTrpLeuLysLysArgThrGlyProAlaAlaThrThr    675680685    LeuArgGlnGlnTyrGlyValArgGlyTyrProThrIleLysPhePhe    690695700    LysAsnGlyAspThrAlaSerProLysGluTyrThrAlaGlyArgGlu    705710715720    AlaAspAspIleValAsnTrpLeuLysLysArgThrGlyProAlaAla    725730735    ThrThrLeuAlaGlnGlnTyrGlyValArgGlyTyrProThrIleLys    740745750    PhePheArgAsnGlyAspLysAlaAlaProArgGluTyrThrAlaGly    755760765    ArgGluAlaAspAspIleValSerTrpLeuLysLysArgThrGlyPro    770775780    AlaAlaThrThrLeuThrMetGluHisAsnIleProGlyPheProSer    785790795800    LeuLysIlePheLysAsnSerAspValAsnAsnSerIleAspTyrGlu    805810815    GlyProArgThrAlaGluAlaValGlnPheMetIleLysGlnSerGln    820825830    ProAlaValAlaValValAlaAspThrAlaAlaAlaGluSerLeuVal    835840845    AspSerSerGluValThrValIleGlyPhePheLysAspAlaGlySer    850855860    AspSerAlaLysGlnPheLeuLeuAlaAlaGluAlaValAspAspIle    865870875880    ProPheGlyIleThrSerAsnSerAspAspGlyAlaAlaAlaGluAla    885890895    LeuValGluSerSerGluValAlaValIleGlyPhePheLysAspMet    900905910    GluSerAspSerAlaLysGlnPhePheLeuAlaAlaGluValIleAsp    915920925    AspIleProPheGlyIleThrSerAsnSerAspAspGlyAlaAlaAla    930935940    GluSerLeuValGluSerSerGluValAlaValIleGlyPhePheLys    945950955960    AspValGluSerAspSerAlaLysGlnPheLeuGlnAlaAlaGluAla    965970975    IleAspAspIleProPheGlyIleThrSerAsnSerAspAspSerAla    980985990    AlaAlaGluSerLeuValGluSerSerGluValAlaValIleGlyPhe    99510001005    PheLysAspValGluSerAspAlaAlaLysGlnPheLeuLeuAlaAla    101010151020    GluAlaThrAspAspIleProPheGlyLeuThrAlaSerSerAspAsp    1025103010351040    AlaAlaAlaAlaGluThrLeuValAspSerSerGluValValValIle    104510501055    GlyPhePheLysAspValThrSerAspAlaAlaLysGluPheLeuLeu    106010651070    AlaAlaGluSerValAspAspIleProPheGlyIleSerSerSerAla    107510801085    AspAspLeuProAlaTyrLeuAlaAsnGluThrPheValThrProVal    109010951100    IleValGlnSerGlyLysIleAspAlaAspPheAsnAlaThrPheTyr    1105111011151120    SerMetAlaAsnLysHisPheAsnAspTyrAspPheValSerAlaGlu    112511301135    AsnAlaAspValPheSerLysTyrGlnLeuAspLysAspGlyValVal    114011451150    LeuPheLysLysPheAspGluGlyArgAsnAsnPheGluGlyGluIle    115511601165    ThrLysGluLysLeuLeuAspPheIleLysHisAsnGlnLeuProLeu    117011751180    ValIleValPheSerLysTyrGlnLeuAspLysAspGlyValValLeu    1185119011951200    PheLysLysPheAspGluGlyArgAsnAsnPheGluGlyGluValThr    120512101215    LysGluLysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuVal    122012251230    IleValPheSerLysTyrGlnLeuAspLysAspGlyValValLeuPhe    123512401245    LysLysPheAspGluGlyArgAsnAsnPheGluGlyGluValThrLys    125012551260    GluAsnLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIle    1265127012751280    ValPheSerArgTyrGlnValHisGlnAspGlyValValLeuPheLys    128512901295    LysPheAspGluGlyArgAsnAsnPheGluGlyGluValThrLysGlu    130013051310    LysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIleVal    131513201325    PheSerLysTyrGlnLeuSerGlnAspGlyValValLeuPheLysLys    133013351340    PheAspGluGlyArgAsnAsnPheGluGlyAspLeuThrLysAspAsn    1345135013551360    LeuLeuAsnPheIleLysSerAsnGlnLeuProLeuValIleAspAsp    136513701375    PheLysLeuSerIleTyrLeuProSerAlaMetAspGluProValVal    138013851390    TyrAsnGlyLysLysAlaAspIleAlaAspAlaAspValPheGluLys    139514001405    TrpLeuGlnValGluAlaLeuProTyrPheGlyGluPheThrGluGln    141014151420    ThrAlaProLysIlePheGlyGlyGluIleLysThrHisIleLeuLeu    1425143014351440    PheLeuProLysSerValSerAspTyrAspGlyLysLeuSerAsnPhe    144514501455    LysLysAlaAlaGluGlyPheLysGlyLysIleGluPheThrGluGln    146014651470    ThrAlaProLysIlePheGlyGlyGluIleLysThrHisIleLeuLeu    147514801485    PheLeuProLysSerValSerAspTyrGluGlyLysLeuSerAsnPhe    149014951500    LysLysAlaAlaGluSerPheLysGlyLysIleGluPheThrGluGln    1505151015151520    ThrAlaProLysIlePheGlyGlyGluIleLysThrHisIleLeuLeu    152515301535    PheLeuProLysSerValSerAspTyrAspGlyLysLeuSerAsnPhe    154015451550    LysThrAlaAlaGluSerPheLysGlyLysIleGluPheThrGluGln    155515601565    ThrAlaProLysIlePheGlyGlyGluIleLysThrHisIleLeuLeu    157015751580    PheLeuProArgSerAlaAlaAspHisAspGlyLysLeuSerGlyPhe    1585159015951600    LysGlnAlaAlaGluGlyPheLysGlyLysIleGluPheThrGluGln    160516101615    ThrAlaProLysIlePheGlyGlyGluIleLysThrHisIleLeuLeu    162016251630    PheLeuProLysSerValSerAspTyrGluGlyLysLeuAspAsnPhe    163516401645    LysThrAlaAlaGlyAsnPheLysGlyLysIleGluIleAspGlySer    165016551660    ValPheAlaGlnTyrValGluSerGlyLeuProLeuGlyTyrLeuPhe    1665167016751680    TyrAsnAspGluGluGluLeuGluGluTyrLysProLeuPheThrGlu    168516901695    LeuAlaLysLysAsnArgGlyLeuPheIlePheIleAspSerAspHis    170017051710    ThrAspAsnGlnArgIleLeuGluPhePheGlyLeuLysLysGluGlu    171517201725    CysProAlaValArgLeuIleThrLeuGluGluGluMetThrLysTyr    173017351740    LeuPheIlePheIleAspSerAspHisThrAspAsnGlnArgIleLeu    1745175017551760    GluPhePheGlyLeuLysLysGluGluCysProAlaValArgLeuIle    176517701775    ThrLeuGluGluGluMetThrLysTyrLeuPheIlePheIleAspSer    178017851790    AspHisThrAspAsnGlnArgIleLeuGluPhePheGlyLeuLysLys    179518001805    GluGluCysProAlaValArgLeuIleThrLeuGluGluGluMetThr    181018151820    LysTyrLeuPheIlePheIleAspSerAspHisAlaAspAsnGlnArg    1825183018351840    IleLeuGluPhePheGlyLeuLysLysGluGluCysProAlaValArg    184518501855    LeuIleThrLeuGluGluGluMetThrLysTyrLeuPheIlePheIle    186018651870    AspSerAspHisSerAspAsnGlnArgIleLeuGluPhePheGlyLeu    187518801885    LysLysGluGluCysProAlaValArgLeuIleThrLeuGluGluGlu    189018951900    MetThrLysTyrLeuMetAsnPheValSerIleAspAlaArgLysPhe    1905191019151920    GlyArgHisAlaGlyAsnLeuAsnMetLysGluGlnPheProLeuPhe    192519301935    AlaIleHisAspMetThrGluAspLeuLysTyrGlyLeuProGlnLeu    194019451950    SerGluGluAlaPheLysProGluSerAspGluLeuThrAlaGluLys    195519601965    IleThrGlnPheCysHisHisPheLeuGluGlyLysIleLysProHis    197019751980    LeuMetSerGlnGluLeuProGluAspTrpAspLysGlnProValLys    1985199019952000    ValLeuValGlyLysLysProGluSerAspGluLeuThrAlaGluLys    200520102015    IleThrGluPheCysHisArgPheLeuGluGlyLysIleLysProHis    202020252030    LeuMetSerGlnGluLeuProAspAspTrpAspLysGlnProValLys    203520402045    ValLeuValGlyLysLysProGluSerGluGluLeuThrAlaGluArg    205020552060    IleThrGluPheCysHisArgPheLeuGluGlyLysIleLysProHis    2065207020752080    LeuMetSerGlnGluArgAlaAspGlyAspTrpAspLysGlnProVal    208520902095    LysValProValGlyLysLysProGluSerAspGluLeuThrAlaGlu    210021052110    GlyIleThrGluPheCysGlnArgPheLeuGluGlyLysIleLysPro    211521202125    HisLeuMetSerGlnGluLeuProAspGluAspTrpAspArgGlnPro    213021352140    ValLysValLeuValGlyLysLysProGluSerAspAspLeuThrAla    2145215021552160    AspLysIleLysGluPheCysAsnLysPheLeuGluGlyLysIleLys    216521702175    ProHisLeuMetSerGlnAspLeuProGluAspTrpAspLysGlnPro    218021852190    ValLysValLeuValGlyLysAspGluLeuSerAspLysIleValLeu    219522002205    GluSerLysAlaIleGluSerLeuAsxLysAspPheLeuLysGlyAsp    221022152220    AlaSerProIleValLysSerGlnGluIlePheGluAsnGlnAspSer    2225223022352240    SerValPheGlnLeuValGlyLysAsnPheGluGluValAlaPheAsp    224522502255    GluLysLysAsnValPheValGluPheTyrAlaProTrpCysGlyHis    226022652270    CysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyrLys    227522802285    AspHisGluAsnIleIleIleAlaLysAsnPheGluGluValAlaPhe    229022952300    AspGluLysLysAsnValPheValGluPheTyrAlaProTrpCysGly    2305231023152320    HisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyr    232523302335    LysAspHisGluAsnIleIleIleAlaLysAsnPheGluAspValAla    234023452350    PheAspGluLysLysAsnValPheValGluPheTyrAlaProTrpCys    235523602365    GlyHisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThr    237023752380    TyrLysAspHisGluAsnIleIleIleAlaLysAsnPheGluGluVal    2385239023952400    AlaPheAspGluLysLysAsnValPheValGluPheTyrAlaProTrp    240524102415    CysGlyHisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGlu    242024252430    ThrTyrLysGluHisGlnAspIleValIleAlaLysAsnPheGluGlu    243524402445    ValAlaPheAspGluAsnLysAsnValPheValGluPheTyrAlaPro    245024552460    TrpCysGlyHisCysLysGlnLeuAlaProAlaTrpAspLysLeuGly    2465247024752480    ProThrTyrArgAspHisGluAsnIleValIleAlaLysAsnHisAsp    248524902495    GluIleValAsnAspProLysLysAspValLeuValLeuTyrTyrAla    250025052510    ProTrpCysGlyHisCysLysArgLeuAlaProThrTyrGlnGluLeu    251525202525    AlaAspThrTyrAlaAsnAlaThrSerAspValLeuIleAlaLysMet    253025352540    AspSerThrAlaAsnGluValGluAlaValLysValHisSerPhePro    2545255025552560    ThrLeuLysPhePheProAlaSerAlaAspArgThrValIleAspTyr    256525702575    AsnGlyGluArgThrLeuAspGlyPheLysLysPheLeuGluSerGly    258025852590    GlyMetAspSerThrAlaAsnGluValGluAlaValLysValHisSer    259526002605    PheProThrLeuLysPhePheProAlaSerAlaAspArgThrValIle    261026152620    AspTyrAsnGlyGluArgThrLeuAspGlyPheLysLysPheLeuGlu    2625263026352640    SerGlyGlyMetAspSerThrAlaAsnGluValGluAlaValLysVal    264526502655    HisSerPheProThrLeuLysPhePheProAlaSerAlaAspArgThr    266026652670    ValIleAspTyrAsnGlyGluArgThrLeuAspGlyPheLysLysPhe    267526802685    LeuGluSerGlyGlyMetAspSerThrAlaAsnGluValGluAlaVal    269026952700    LysValHisSerPheProThrLeuLysPhePheProAlaGlyProGly    2705271027152720    ArgThrValIleAspTyrAsnGlyGluArgThrLeuAspGlyPheLys    272527302735    LysPheLeuGluSerGlyGlyMetAspSerThrAlaAsnGluValGlu    274027452750    AlaValLysValHisSerPheProThrLeuLysPhePheProAlaGly    275527602765    SerGlyArgAsnValIleAspTyrAsnGlyGluArgThrLeuGluGly    277027752780    PheLysLysPheLeuGluSerGlyGlyLeuAspHisThrGluAsnAsp    2785279027952800    ValArgGlyValValIleGluGlyTyrProThrIleValLeuTyrPro    280528102815    GlyGlyLysLysSerGluSerValValTyrGlnGlySerArgSerLeu    282028252830    AspSerLeuPheAspPheIleLysGluAsnGlyGlnAspGlyAlaGly    283528402845    AspAsnAspAspLeuAspLeuGluGluAlaLeuGluProAspMetGlu    285028552860    GluAspAspAspGlnLysAlaValLysAspGluLeuGlnAspGlyAla    2865287028752880    GlyAspAspAspAspLeuGluAspLeuGluGluAlaGluGluProAsp    288528902895    LeuGluGluAspAspAspGlnLysAlaValLysAspGluLeuGlnAsp    290029052910    GlyAlaGlyAspAspAspAspLeuGluAspLeuGluGluAlaGluGlu    291529202925    ProAspMetGluGluAspAspAspGlnLysAlaValLysAspGluLeu    293029352940    GlnAspGlyAlaGlyAspGluAspGlyLeuGluAspLeuGluGluAla    2945295029552960    GluGluProAspLeuGluGluAspAspAspGlnLysAlaValArgAsp    296529702975    GluLeuGlnAspGlyAlaAlaAlaAspAspAspLeuGluAspLeuGlu    298029852990    ThrAspGluGluThrAspLeuGluGluGlyAspAspAspGluGlnLys    299530003005    IleGlnLysAspGluLeuHisPheAspValAspGlyLysAlaLeuTyr    301030153020    GluGluAlaGlnGluLysAlaAlaGluGluAlaAspAlaAspAlaGlu    3025303030353040    LeuAlaAspGluGluAspAlaIleHisAspGluLeu    30453050    (2) INFORMATION FOR SEQ ID NO:27:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 509 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:    MetLeuSerArgAlaLeuLeuCysLeuAlaLeuAlaTrpAlaAlaArg    151015    ValGlyAlaAspAlaLeuGluGluGluAspAsnValLeuValLeuLys    202530    LysSerAsnPheAlaGluAlaLeuAlaAlaHisAsnTyrLeuLeuVal    354045    GluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGlu    505560    TyrAlaLysAlaAlaAlaLysLeuLysAlaGluGlySerGluIleArg    65707580    LeuAlaLysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyr    859095    GlyValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThr    100105110    AlaSerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleVal    115120125    AsnTrpLeuLysLysArgThrGlyProAlaAlaThrThrLeuSerAsp    130135140    ThrAlaAlaAlaGluSerLeuValAspSerSerGluValThrValIle    145150155160    GlyPhePheLysAspAlaGlySerAspSerAlaLysGlnPheLeuLeu    165170175    AlaAlaGluAlaValAspAspIleProPheGlyIleThrSerAsnSer    180185190    AspValPheSerLysTyrGlnLeuAspLysAspGlyValValLeuPhe    195200205    LysLysPheAspGluGlyArgAsnAsnPheGluGlyGluIleThrLys    210215220    GluLysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIle    225230235240    GluPheThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLys    245250255    ThrHisIleLeuLeuPheLeuProLysSerValSerAspTyrAspGly    260265270    LysLeuSerAsnPheLysLysAlaAlaGluGlyPheLysGlyLysIle    275280285    LeuPheIlePheIleAspSerAspHisThrAspAsnGlnArgIleLeu    290295300    GluPhePheGlyLeuLysLysGluGluCysProAlaValArgLeuIle    305310315320    ThrLeuGluGluGluMetThrLysTyrLysProGluSerAspGluLeu    325330335    ThrAlaGluLysIleThrGlnPheCysHisHisPheLeuGluGlyLys    340345350    IleLysProHisLeuMetSerGlnGluLeuProGluAspTrpAspLys    355360365    GlnProValLysValLeuValGlyLysAsnPheGluGluValAlaPhe    370375380    AspGluLysLysAsnValPheValGluPheTyrAlaProTrpCysGly    385390395400    HisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyr    405410415    LysAspHisGluAsnIleIleIleAlaLysMetAspSerThrAlaAsn    420425430    GluValGluAlaValLysValHisSerPheProThrLeuLysPhePhe    435440445    ProAlaSerAlaAspArgThrValIleAspTyrAsnGlyGluArgThr    450455460    LeuAspGlyPheLysLysPheLeuGluSerGlyGlyGlnAspGlyAla    465470475480    GlyAspAsnAspAspLeuAspLeuGluGluAlaLeuGluProAspMet    485490495    GluGluAspAspAspGlnLysAlaValLysAspGluLeu    500505    (2) INFORMATION FOR SEQ ID NO:28:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 510 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:    MetLeuArgArgAlaLeuLeuCysLeuAlaLeuThrAlaLeuPheArg    151015    AlaGlyAlaGlyAlaProAspGluGluAspHisValLeuValLeuHis    202530    LysGlyAsnPheAspGluAlaLeuAlaAlaHisLysTyrLeuLeuVal    354045    GluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGlu    505560    TyrAlaLysAlaAlaGlyLysLeuLysAlaGluGlySerGluIleArg    65707580    LeuAlaLysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyr    859095    GlyValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThr    100105110    AlaSerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleVal    115120125    AsnTrpLeuLysLysArgThrGlyProAlaAlaSerThrLeuSerAsp    130135140    GlyAlaAlaAlaGluAlaLeuValGluSerSerGluValAlaValIle    145150155160    GlyPhePheLysAspMetGluSerAspSerAlaLysGlnPhePheLeu    165170175    AlaAlaGluValIleAspAspIleProPheGlyIleThrSerAsnSer    180185190    AspValPheSerLysTyrGlnLeuAspLysAspGlyValValLeuPhe    195200205    LysLysPheAspGluGlyArgAsnAsnPheGluGlyGluValThrLys    210215220    GluLysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIle    225230235240    GluPheThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLys    245250255    ThrHisIleLeuLeuPheLeuProLysSerValSerAspTyrGluGly    260265270    LysLeuSerAsnPheLysLysAlaAlaGluSerPheLysGlyLysIle    275280285    LeuPheIlePheIleAspSerAspHisThrAspAsnGlnArgIleLeu    290295300    GluPhePheGlyLeuLysLysGluGluCysProAlaValArgLeuIle    305310315320    ThrLeuGluGluGluMetThrLysTyrLysProGluSerAspGluLeu    325330335    ThrAlaGluLysIleThrGluPheCysHisArgPheLeuGluGlyLys    340345350    IleLysProHisLeuMetSerGlnGluLeuProAspAspTrpAspLys    355360365    GlnProValLysValLeuValGlyLysAsnPheGluGluValAlaPhe    370375380    AspGluLysLysAsnValPheValGluPheTyrAlaProTrpCysGly    385390395400    HisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyr    405410415    LysAspHisGluAsnIleIleIleAlaLysMetAspSerThrAlaAsn    420425430    GluValGluAlaValLysValHisSerPheProThrLeuLysPhePhe    435440445    ProAlaSerAlaAspArgThrValIleAspTyrAsnGlyGluArgThr    450455460    LeuAspGlyPheLysLysPheLeuGluSerGlyGlyGlnAspGlyAla    465470475480    GlyAspAspAspAspLeuGluAspLeuGluGluAlaGluGluProAsp    485490495    LeuGluGluAspAspAspGlnLysAlaValLysAspGluLeu    500505510    (2) INFORMATION FOR SEQ ID NO:29:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 509 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:    MetLeuArgArgAlaLeuLeuCysLeuAlaValAlaAlaLeuValArg    151015    AlaAspAlaProGluGluGluAspHisValLeuValLeuArgLysSer    202530    AsnPheAlaGluAlaLeuAlaAlaHisLysTyrLeuLeuValGluPhe    354045    TyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyrAla    505560    LysAlaAlaGlyLysLeuLysAlaGluGlySerGluIleArgLeuAla    65707580    LysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyrGlyVal    859095    ArgGlyTyrProThrIleLysPhePheArgAsnGlyAspThrAlaSer    100105110    ProLysGluTyrThrAlaGlyArgGluAlaAspAspIleValAsnTrp    115120125    LeuLysLysArgThrGlyProAlaAlaThrThrLeuArgAspGlyAla    130135140    AlaAlaGluSerLeuValGluSerSerGluValAlaValIleGlyPhe    145150155160    PheLysAspValGluSerAspSerAlaLysGlnPheLeuGlnAlaAla    165170175    GluAlaIleAspAspIleProPheGlyIleThrSerAsnSerAspVal    180185190    PheSerLysTyrGlnLeuAspLysAspGlyValValLeuPheLysLys    195200205    PheAspGluGlyArgAsnAsnPheGluGlyGluValThrLysGluAsn    210215220    LeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIleGluPhe    225230235240    ThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLysThrHis    245250255    IleLeuLeuPheLeuProLysSerValSerAspTyrAspGlyLysLeu    260265270    SerAsnPheLysThrAlaAlaGluSerPheLysGlyLysIleLeuPhe    275280285    IlePheIleAspSerAspHisThrAspAsnGlnArgIleLeuGluPhe    290295300    PheGlyLeuLysLysGluGluCysProAlaValArgLeuIleThrLeu    305310315320    GluGluGluMetThrLysTyrLysProGluSerGluGluLeuThrAla    325330335    GluArgIleThrGluPheCysHisArgPheLeuGluGlyLysIleLys    340345350    ProHisLeuMetSerGlnGluArgAlaAspGlyAspTrpAspLysGln    355360365    ProValLysValProValGlyLysAsnPheGluAspValAlaPheAsp    370375380    GluLysLysAsnValPheValGluPheTyrAlaProTrpCysGlyHis    385390395400    CysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyrLys    405410415    AspHisGluAsnIleIleIleAlaLysMetAspSerThrAlaAsnGlu    420425430    ValGluAlaValLysValHisSerPheProThrLeuLysPhePhePro    435440445    AlaSerAlaAspArgThrValIleAspTyrAsnGlyGluArgThrLeu    450455460    AspGlyPheLysLysPheLeuGluSerGlyGlyGlnAspGlyAlaGly    465470475480    AspAspAspAspLeuGluAspLeuGluGluAlaGluGluProAspMet    485490495    GluGluAspAspAspGlnLysAlaValLysAspGluLeu    500505    (2) INFORMATION FOR SEQ ID NO:30:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 510 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:    MetLeuArgArgAlaValLeuCysLeuAlaLeuAlaValThrAlaGly    151015    TrpAlaTrpAlaAlaGluGluGluAspAsnValLeuValLeuLysSer    202530    SerAsnPheAlaGluGluLeuAlaAlaHisLysTyrLeuLeuValGlu    354045    PheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyr    505560    AlaLysAlaAlaGlyLysLeuLysAlaGluGlySerAspIleArgLeu    65707580    AlaLysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyrGly    859095    ValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThrAla    100105110    SerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleValAsn    115120125    TrpLeuLysLysArgThrGlyProAlaAlaThrThrLeuAlaAspSer    130135140    AlaAlaAlaGluSerLeuValGluSerSerGluValAlaValIleGly    145150155160    PhePheLysAspValGluSerAspAlaAlaLysGlnPheLeuLeuAla    165170175    AlaGluAlaThrAspAspIleProPheGlyLeuThrAlaSerSerAsp    180185190    ValPheSerArgTyrGlnValHisGlnAspGlyValValLeuPheLys    195200205    LysPheAspGluGlyArgAsnAsnPheGluGlyGluValThrLysGlu    210215220    LysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIleGlu    225230235240    PheThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLysThr    245250255    HisIleLeuLeuPheLeuProArgSerAlaAlaAspHisAspGlyLys    260265270    LeuSerGlyPheLysGlnAlaAlaGluGlyPheLysGlyLysIleLeu    275280285    PheIlePheIleAspSerAspHisAlaAspAsnGlnArgIleLeuGlu    290295300    PhePheGlyLeuLysLysGluGluCysProAlaValArgLeuIleThr    305310315320    LeuGluGluGluMetThrLysTyrLysProGluSerAspGluLeuThr    325330335    AlaGluGlyIleThrGluPheCysGlnArgPheLeuGluGlyLysIle    340345350    LysProHisLeuMetSerGlnGluLeuProAspGluAspTrpAspArg    355360365    GlnProValLysValLeuValGlyLysAsnPheGluGluValAlaPhe    370375380    AspGluLysLysAsnValPheValGluPheTyrAlaProTrpCysGly    385390395400    HisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyr    405410415    LysGluHisGlnAspIleValIleAlaLysMetAspSerThrAlaAsn    420425430    GluValGluAlaValLysValHisSerPheProThrLeuLysPhePhe    435440445    ProAlaGlyProGlyArgThrValIleAspTyrAsnGlyGluArgThr    450455460    LeuAspGlyPheLysLysPheLeuGluSerGlyGlyGlnAspGlyAla    465470475480    GlyAspGluAspGlyLeuGluAspLeuGluGluAlaGluGluProAsp    485490495    LeuGluGluAspAspAspGlnLysAlaValArgAspGluLeu    500505510    (2) INFORMATION FOR SEQ ID NO:31:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 493 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:31:    GluProLeuGluGluGluAspGlyValLeuValLeuArgAlaAlaAsn    151015    PheGluGlnAlaLeuAlaAlaHisArgHisLeuLeuValGluPheTyr    202530    AlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyrAlaLys    354045    AlaAlaAlaGlnLeuLysAlaGluGlySerGluIleArgLeuAlaLys    505560    ValAspAlaThrGluGluAlaGluLeuAlaGlnGlnPheGlyValArg    65707580    GlyTyrProThrIleLysPhePheArgAsnGlyAspLysAlaAlaPro    859095    ArgGluTyrThrAlaGlyArgGluAlaAspAspIleValSerTrpLeu    100105110    LysLysArgThrGlyProAlaAlaThrThrLeuThrAspAlaAlaAla    115120125    AlaGluThrLeuValAspSerSerGluValValValIleGlyPhePhe    130135140    LysAspValThrSerAspAlaAlaLysGluPheLeuLeuAlaAlaGlu    145150155160    SerValAspAspIleProPheGlyIleSerSerSerAlaAspValPhe    165170175    SerLysTyrGlnLeuSerGlnAspGlyValValLeuPheLysLysPhe    180185190    AspGluGlyArgAsnAsnPheGluGlyAspLeuThrLysAspAsnLeu    195200205    LeuAsnPheIleLysSerAsnGlnLeuProLeuValIleGluPheThr    210215220    GluGlnThrAlaProLysIlePheGlyGlyGluIleLysThrHisIle    225230235240    LeuLeuPheLeuProLysSerValSerAspTyrGluGlyLysLeuAsp    245250255    AsnPheLysThrAlaAlaGlyAsnPheLysGlyLysIleLeuPheIle    260265270    PheIleAspSerAspHisSerAspAsnGlnArgIleLeuGluPhePhe    275280285    GlyLeuLysLysGluGluCysProAlaValArgLeuIleThrLeuGlu    290295300    GluGluMetThrLysTyrLysProGluSerAspAspLeuThrAlaAsp    305310315320    LysIleLysGluPheCysAsnLysPheLeuGluGlyLysIleLysPro    325330335    HisLeuMetSerGlnAspLeuProGluAspTrpAspLysGlnProVal    340345350    LysValLeuValGlyLysAsnPheGluGluValAlaPheAspGluAsn    355360365    LysAsnValPheValGluPheTyrAlaProTrpCysGlyHisCysLys    370375380    GlnLeuAlaProAlaTrpAspLysLeuGlyProThrTyrArgAspHis    385390395400    GluAsnIleValIleAlaLysMetAspSerThrAlaAsnGluValGlu    405410415    AlaValLysIleHisSerPheProThrLeuLysPhePheProAlaGly    420425430    SerGlyArgAsnValIleAspTyrAsnGlyGluArgThrLeuGluGly    435440445    PheLysLysPheLeuGluSerGlyGlyGlnAspGlyAlaAlaAlaAsp    450455460    AspAspLeuGluAspLeuGluThrAspGluGluThrAspLeuGluGlu    465470475480    GlyAspAspAspGluGlnLysIleGlnLysAspGluLeu    485490    (2) INFORMATION FOR SEQ ID NO:32:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 521 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:32:    MetLysPheSerAlaGlyAlaValLeuSerTrpSerSerLeuLeuLeu    151015    AlaSerSerValPheAlaGlnGlnGluAlaValAlaProGluAspSer    202530    AlaValValLysLeuAlaThrAspSerPheAsnGluTyrIleGlnSer    354045    HisAspLeuValLeuAlaGluPhePheAlaProTrpCysGlyHisCys    505560    LysAsnMetAlaProGluTyrValLysAlaAlaGluThrLeuValGlu    65707580    LysAsnIleThrLeuAlaGlnIleAspCysThrGluAsnGlnAspLeu    859095    CysMetGluHisAsnIleProGlyPheProSerLeuLysIlePheLys    100105110    AsnSerAspValAsnAsnSerIleAspTyrGluGlyProArgThrAla    115120125    GluAlaValGlnPheMetIleLysGlnSerGlnProAlaValAlaVal    130135140    ValAlaAspLeuProAlaTyrLeuAlaAsnGluThrPheValThrPro    145150155160    ValIleValGlnSerGlyLysIleAspAlaAspPheAsnAlaThrPhe    165170175    TyrSerMetAlaAsnLysHisPheAsnAspTyrAspPheValSerAla    180185190    GluAsnAlaAspAspAspPheLysLeuSerIleTyrLeuProSerAla    195200205    MetAspGluProValValTyrAsnGlyLysLysAlaAspIleAlaAsp    210215220    AlaAspValPheGluLysTrpLeuGlnValGluAlaLeuProTyrPhe    225230235240    GlyGluIleAspGlySerValPheAlaGlnTyrValGluSerGlyLeu    245250255    ProLeuGlyTyrLeuPheTyrAsnAspGluGluGluLeuGluGluTyr    260265270    LysProLeuPheThrGluLeuAlaLysLysAsnArgGlyLeuMetAsn    275280285    PheValSerIleAspAlaArgLysPheGlyArgHisAlaGlyAsnLeu    290295300    AsnMetLysGluGlnPheProLeuPheAlaIleHisAspMetThrGlu    305310315320    AspLeuLysTyrGlyLeuProGlnLeuSerGluGluAlaPheAspGlu    325330335    LeuSerAspLysIleValLeuGluSerLysAlaIleGluSerLeuAsx    340345350    LysAspPheLeuLysGlyAspAlaSerProIleValLysSerGlnGlu    355360365    IlePheGluAsnGlnAspSerSerValPheGlnLeuValGlyLysAsn    370375380    HisAspGluIleValAsnAspProLysLysAspValLeuValLeuTyr    385390395400    TyrAlaProTrpCysGlyHisCysLysArgLeuAlaProThrTyrGln    405410415    GluLeuAlaAspThrTyrAlaAsnAlaThrSerAspValLeuIleAla    420425430    LysLeuAspHisThrGluAsnAspValArgGlyValValIleGluGly    435440445    TyrProThrIleValLeuTyrProGlyGlyLysLysSerGluSerVal    450455460    ValTyrGlnGlySerArgSerLeuAspSerLeuPheAspPheIleLys    465470475480    GluAsnGlyHisPheAspValAspGlyLysAlaLeuTyrGluGluAla    485490495    GlnGluLysAlaAlaGluGluAlaAspAlaAspAlaGluLeuAlaAsp    500505510    GluGluAspAlaIleHisAspGluLeu    515520    (2) INFORMATION FOR SEQ ID NO:33:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 512 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:33:    MetAlaLysAsnValAlaIlePheGlyLeuLeuPheSerLeuLeuLeu    151015    LeuValProSerGlnIlePheAlaGluGluSerSerThrAspAlaLys    202530    GluPheValLeuThrLeuAspAsnThrAsnPheHisAspThrValLys    354045    LysHisAspPheIleValValGluPheTyrAlaProTrpCysGlyHis    505560    CysLysLysLeuAlaProGluTyrGluLysAlaAlaSerIleLeuSer    65707580    ThrHisGluProProValValLeuAlaLysValAspAlaAsnGluGlu    859095    HisAsnLysAspLeuAlaSerGluAsnAspValLysGlyPheProThr    100105110    IleLysIlePheArgAsnGlyGlyLysAsnIleGlnGluTyrLysGly    115120125    ProArgGluAlaGluGlyIleValGluTyrLeuLysLysGlnSerGly    130135140    ProAlaSerThrGluIleLysSerAlaAspAspAlaThrAlaPheVal    145150155160    GlyAspAsnLysValValIleValGlyValPheProLysPheSerGly    165170175    GluGluTyrAspAsnPheIleAlaLeuAlaGluLysLeuArgSerAsp    180185190    TyrAspPheAlaHisThrLeuAsnAlaLysHisLeuProLysGlyAsp    195200205    SerSerValSerGlyProValValArgLeuPheLysProPheAspGlu    210215220    LeuPheValAspSerLysAspPheAsnValGluAlaLeuGluLysPhe    225230235240    IleGluGluSerSerThrProIleValThrValPheAsnAsnGluPro    245250255    SerAsnHisProPheValValLysPhePheAsnSerProAsnAlaLys    260265270    AlaMetLeuPheIleAsnPheThrThrGluGlyAlaGluSerPheLys    275280285    ThrLysTyrHisGluValAlaGluGlnTyrLysGlnGlnGlyValSer    290295300    PheLeuValGlyAspValGluSerSerGlnGlyAlaPheGlnTyrPhe    305310315320    GlyLeuLysGluGluGlnValProLeuIleIleIleGlnHisAsnAsp    325330335    GlyLysLysPhePheLysProAsnLeuGluLeuAspGlnLeuProThr    340345350    TrpLeuLysAlaTyrLysAspGlyLysValGluProPheValLysSer    355360365    GluProIleProGluThrAsnAsnGluProValLysValValValGly    370375380    GlnThrLeuGluAspValValPheLysSerGlyLysAsnValLeuIle    385390395400    GluPheTyrAlaProTrpCysGlyHisCysLysGlnLeuAlaProIle    405410415    LeuAspGluValAlaValSerPheGlnSerAspAlaAspValValIle    420425430    AlaLysLeuAspAlaThrAlaAsnAspIleProThrAspThrPheAsp    435440445    ValGlnGlyTyrProThrLeuTyrPheArgSerAlaSerGlyLysLeu    450455460    SerGlnTyrAspGlyGlyArgThrLysGluAspIleIleGluPheIle    465470475480    GluLysAsnLysAspLysThrGlyAlaAlaHisGlnGluValGluGln    485490495    ProLysAlaAlaAlaGlnProGluAlaGluGlnProLysAspGluLeu    500505510    (2) INFORMATION FOR SEQ ID NO:34:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 515 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:    MetArgThrPheAlaProTrpIleLeuSerLeuLeuGlyAlaSerAla    151015    ValAlaSerAlaAlaAspAlaThrAlaGluAlaProSerAspValVal    202530    SerLeuThrGlyAspThrPheGluThrPheValLysGluHisAspLeu    354045    ValLeuAlaGluPhePheAlaProTrpCysGlyHisCysLysAlaLeu    505560    AlaProLysTyrGluGlnAlaAlaThrGluLeuLysGluLysAsnIle    65707580    ProLeuValLysValAspCysThrGluGluGluAlaLeuCysArgAsp    859095    GlnGlyValGluGlyTyrProThrLeuLysIlePheArgGlyLeuAsp    100105110    AlaValLysProTyrGlnGlyAlaArgGlnThrGluAlaIleValSer    115120125    TyrMetValLysGlnSerLeuProAlaValSerProValThrProGlu    130135140    AsnLeuGluGluIleLysThrMetAspLysIleValValIleGlyTyr    145150155160    IleAlaSerAspAspGlnThrAlaAsnAspIlePheThrThrPheAla    165170175    GluSerGlnArgAspAsnTyrLeuPheAlaAlaThrSerAspAlaSer    180185190    IleAlaLysAlaGluGlyValLysGlnProSerIleValLeuTyrLys    195200205    AspPheAspGluLysLysAlaThrTyrAspGlyGluIleGluGlnAsp    210215220    AlaLeuLeuSerTrpValLysThrAlaSerThrProLeuValGlyGlu    225230235240    LeuGlyProGluThrTyrSerGlyTyrIleThrAlaGlyIleProLeu    245250255    AlaTyrIlePheAlaGluThrLysGluGluArgGluGlnPheThrGlu    260265270    GluPheLysPheIleAlaGluLysHisLysGlySerIleAsnIleVal    275280285    ThrIleAspAlaLysLeuTyrGlyAlaHisAlaGlyAsnLeuAsnLeu    290295300    AspProSerLysPheProAlaPheAlaIleGlnAspProGluLysAsn    305310315320    AlaLysTyrProTyrAspGlnSerLysGluValLysAlaLysAspIle    325330335    GlyLysPheIleGlnAspValLeuAspAspLysValGluProSerIle    340345350    LysSerGluAlaIleProGluThrGlnGluGlyProValThrValVal    355360365    ValAlaHisSerTyrLysAspLeuValLeuAspAsnGluLysAspVal    370375380    LeuLeuGluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAla    385390395400    ProLysTyrGluGluLeuAlaSerLeuTyrLysAspIleProGluVal    405410415    ThrIleAlaLysIleAspAlaThrAlaAsnAspValProAspSerIle    420425430    ThrGlyPheProThrIleLysLeuPheAlaAlaGlyAlaLysAspSer    435440445    ProValGluTyrGluGlySerArgThrValGluAspLeuAlaAsnPhe    450455460    ValLysGluAsnGlyLysHisLysValAspAlaLeuGluValAspPro    465470475480    LysLysGluGlnGluSerGlyAspAlaThrGluThrArgAlaAlaSer    485490495    AspGluThrGluThrProAlaAlaThrSerAspAspLysSerGluHis    500505510    AspGluLeu    515    (2) INFORMATION FOR SEQ ID NO:35:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 530 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:35:    MetLysPheSerAlaGlyAlaValLeuSerTrpSerSerLeuLeuLeu    151015    AlaSerSerValPheAlaGlnGlnGluAlaValAlaProGluAspSer    202530    AlaValValLysLeuAlaThrAspSerPheAsnGluTyrIleGlnSer    354045    HisAspLeuValLeuAlaGluPhePheAlaProTrpCysGlyHisCys    505560    LysAsnMetAlaProGluTyrValLysAlaAlaGluThrLeuValGlu    65707580    LysAsnIleThrLeuAlaGlnIleAspCysThrGluAsnGlnAspLeu    859095    CysMetGluHisAsnIleProGlyPheProSerLeuLysIlePheLys    100105110    AsnArgAspValAsnAsnSerIleAspTyrGluGlyProArgThrAla    115120125    GluAlaIleValGlnPheMetIleLysGlnSerGlnProAlaValAla    130135140    ValValAlaAspLeuProAlaTyrLeuAlaAsnGluThrPheValThr    145150155160    ProValIleValGlnSerGlyLysIleAspAlaAspPheAsnAlaThr    165170175    PheTyrSerMetAlaAsnLysHisPheAsnAspTyrAspPheValSer    180185190    AlaGluAsnAlaAspAspAspPheLysLeuSerIleTyrLeuProSer    195200205    AlaMetAspGluProValValTyrAsnGlyLysLysAlaAspIleAla    210215220    AspAlaAspValPheGluLysTrpLeuGlnValGluAlaLeuProTyr    225230235240    PheGlyGluIleAspGlySerValPheAlaGlnTyrValGluSerGly    245250255    LeuProLeuGlyTyrLeuPheTyrAsnAspGluGluGluLeuGluGlu    260265270    TyrLysProLeuPheThrGluLeuAlaLysLysAsnArgGlyLeuMet    275280285    AsnPheValSerIleAspAlaArgLysPheGlyArgHisAlaGlyAsn    290295300    LeuAsnMetLysGluGlnPheProLeuPheAlaIleHisAspMetThr    305310315320    GluAspLeuLysTyrGlyLeuProGlnLeuSerGluGluAlaPheAsp    325330335    GluLeuSerAspLysIleValLeuGluSerLysAlaIleGluSerLeu    340345350    ValLysAspPheLeuLysGlyAspAlaSerProIleValLysSerGln    355360365    GluIlePheGluAsnGlnAspSerSerValPheGlnLeuValGlyLys    370375380    AsnHisAspGluIleValAsnAspProLysLysAspValLeuValLeu    385390395400    TyrTyrAlaProTrpCysGlyHisCysLysArgLeuAlaProThrTyr    405410415    GlnGluLeuAlaAspThrTyrAlaAsnAlaThrSerAspValLeuIle    420425430    AlaLysLeuAspHisThrGluAsnAspValArgGlyValValIleGlu    435440445    GlyTyrProThrIleValLeuTyrProGlyGlyLysLysSerGluSer    450455460    ValValTyrGlnGlySerArgSerLeuAspSerLeuPheAspPheIle    465470475480    LysGluAsnGlyHisPheAspValAspGlyLysAlaLeuTyrGluGlu    485490495    AlaGlnGluLysAlaAlaGluGluAlaGluAlaAspAlaGluAlaGlu    500505510    AlaAspAlaAspAlaGluLeuAlaAspGluGluAspAlaIleHisAsp    515520525    GluLeu    530    (2) INFORMATION FOR SEQ ID NO:36:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 510 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:    MetLeuArgArgAlaLeuLeuCysLeuAlaLeuThrAlaLeuPheArg    151015    AlaGlyAlaGlyAlaProAspGluGluAspHisValLeuValLeuHis    202530    LysGlyAsnPheAspGluAlaLeuAlaAlaHisLysTyrLeuLeuVal    354045    GluPheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGlu    505560    TyrAlaLysAlaAlaGlyLysLeuLysAlaGluGlySerGluIleArg    65707580    LeuAlaLysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyr    859095    GlyValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThr    100105110    AlaSerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleVal    115120125    AsnTrpLeuLysLysArgThrGlyProAlaAlaSerThrLeuSerAsp    130135140    GlyAlaAlaAlaGluAlaLeuValGluSerSerGluValAlaValIle    145150155160    GlyPhePheLysAspMetGluSerAspSerAlaLysGlnPhePheLeu    165170175    AlaAlaGluValIleAspAspIleProPheGlyIleThrSerAsnSer    180185190    AspValPheSerLysTyrGlnLeuAspLysAspGlyValValLeuPhe    195200205    LysLysPheAspGluGlyArgAsnAsnPheGluGlyGluValThrLys    210215220    GluLysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIle    225230235240    GluPheThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLys    245250255    ThrHisIleLeuLeuPheLeuProLysSerValSerAspTyrGluGly    260265270    LysLeuSerAsnPheLysLysAlaAlaGluSerPheLysGlyLysIle    275280285    LeuPheIlePheIleAspSerAspHisThrAspAsnGlnArgIleLeu    290295300    GluPheGluGlyLeuLysLysGluGluCysProAlaValArgLeuIle    305310315320    ThrLeuGluGluGluMetThrLysTyrLysProGluSerAspGluLeu    325330335    ThrAlaGluLysIleThrGluPheCysHisArgPheLeuGluGlyLys    340345350    IleLysProHisLeuMetSerGlnGluLeuProAspAspTrpAspLys    355360365    GlnProValLysValLeuValGlyLysAsnPheGluGluValAlaPhe    370375380    AspGluLysLysAsnValPheValGluPheTyrAlaProTrpCysGly    385390395400    HisCysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyr    405410415    LysAspHisGluAsnIleValIleAlaLysMetAspSerThrAlaAsn    420425430    GluValGluAlaValLysValHisSerPheProThrLeuLysPhePhe    435440445    ProAlaSerAlaAspArgThrValIleAspTyrAsnGlyGluArgThr    450455460    LeuAspGlyPheLysLysPheLeuGluSerGlyGlyGlnAspGlyAla    465470475480    GlyAspAspAspAspLeuGluAspLeuGluGluAlaGluGluProAsp    485490495    LeuGluGluAspAspAspGlnLysAlaValLysAspGluLeu    500505510    (2) INFORMATION FOR SEQ ID NO:37:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 508 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:37:    MetLeuSerArgAlaLeuLeuCysLeuAlaLeuAlaTrpAlaAlaArg    151015    ValGlyAlaAspAlaLeuGluGluGluAspAsnValLeuValLeuLys    202530    LysSerAsnPheAlaGluProAlaAlaHisAsnTyrLeuLeuValGlu    354045    PheTyrAlaProTrpCysGlyHisCysLysAlaLeuAlaProGluTyr    505560    AlaLysAlaAlaAlaLysLeuLysAlaGluGlySerGluIleArgLeu    65707580    AlaLysValAspAlaThrGluGluSerAspLeuAlaGlnGlnTyrGly    859095    ValArgGlyTyrProThrIleLysPhePheLysAsnGlyAspThrAla    100105110    SerProLysGluTyrThrAlaGlyArgGluAlaAspAspIleValAsn    115120125    TrpLeuLysLysArgThrGlyProAlaAlaThrThrLeuSerAspThr    130135140    AlaAlaAlaGluSerLeuValAspSerSerGluValThrValIleGly    145150155160    PhePheLysAspAlaGlySerAspSerAlaLysGlnPheLeuLeuAla    165170175    AlaGluAlaValAspAspIleProPheGlyIleThrSerAsnSerAsp    180185190    ValPheSerLysTyrGlnLeuAspLysAspGlyValValLeuPheLys    195200205    LysPheAspGluGlyArgAsnAsnPheGluGlyGluIleThrLysGlu    210215220    LysLeuLeuAspPheIleLysHisAsnGlnLeuProLeuValIleGlu    225230235240    PheThrGluGlnThrAlaProLysIlePheGlyGlyGluIleLysThr    245250255    HisIleLeuLeuPheLeuProLysSerValSerAspTyrAspGlyLys    260265270    LeuSerAsnPheLysLysAlaAlaGluGlyPheLysGlyLysIleLeu    275280285    PheIlePheIleAspSerAspHisThrAspAsnGlnArgIleLeuGlu    290295300    PhePheGlyLeuLysLysGluGluCysProAlaValArgLeuIleThr    305310315320    LeuGluGluGluMetThrLysTyrLysProGluSerAspGluLeuThr    325330335    AlaGluLysIleThrGlnPheCysHisHisPheLeuGluGlyLysIle    340345350    LysProHisLeuMetSerGlnGluLeuProGluAspTrpAspLysGln    355360365    ProValLysValLeuValGlyLysAsnPheGluGluValAlaPheAsp    370375380    GluLysLysAsnValPheValGluPheTyrAlaProTrpCysGlyHis    385390395400    CysLysGlnLeuAlaProIleTrpAspLysLeuGlyGluThrTyrLys    405410415    AspHisGluAsnIleValIleAlaLysMetAspSerThrAlaAsnGlu    420425430    ValGluAlaValLysValHisSerPheProThrLeuLysPhePhePro    435440445    AlaSerAlaAspArgThrValIleAspTyrAsnGlyGluArgThrLeu    450455460    AspGlyPheLysLysPheLeuGluSerGlyArgGlnAspGlyAlaGly    465470475480    AspAsnAspAspLeuAspLeuGluGluAlaLeuGluProAspMetGlu    485490495    GluAspAspAspGlnLysAlaValLysAspGluLeu    500505    (2) INFORMATION FOR SEQ ID NO:38:    (i) SEQUENCE CHARACTERISTICS:    (A) LENGTH: 638 amino acids    (B) TYPE: amino acid    (C) STRANDEDNESS: single    (D) TOPOLOGY: linear    (ii) MOLECULE TYPE: peptide    (xi) SEQUENCE DESCRIPTION: SEQ ID NO:38:    MetLysLeuArgLysAlaTrpLeuLeuValLeuLeuLeuAlaLeuThr    151015    GlnLeuLeuAlaAlaAlaSerAlaGlyAspAlaGlnGluAspThrSer    202530    AspThrGluAsnAlaThrGluGluGluGluGluGluAspAspAspAsp    354045    LeuGluValLysGluGluAsnGlyValTrpValLeuAsnAspGlyAsn    505560    PheAspAsnPheValAlaAspLysAspThrValLeuLeuGluPheTyr    65707580    AlaProTrpCysGlyHisCysLysGlnPheAlaProGluTyrGluLys    859095    IleAlaSerThrLeuLysAspAsnAspProProIleAlaValAlaLys    100105110    IleAspAlaThrSerAlaSerMetLeuAlaSerLysPheAspValSer    115120125    GlyTyrProThrIleLysIleLeuLysLysGlyGlnAlaValAspTyr    130135140    AspGlySerArgThrGlnGluGluIleValAlaLysValArgGluVal    145150155160    SerGlnProAspTrpThrProProProGluValThrLeuSerLeuThr    165170175    LysAspAsnPheAspAspValValAsnAsnAlaAspIleIleLeuVal    180185190    GluPheTyrAlaProTrpCysGlyHisCysLysLysLeuAlaProGlu    195200205    TyrGluLysAlaAlaLysGluLeuSerLysArgSerProProIlePro    210215220    LeuAlaLysValAspAlaThrGluGlnThrAspLeuAlaLysArgPhe    225230235240    AspValSerGlyTyrProThrLeuLysIlePheArgLysGlyArgPro    245250255    PheAspTyrAsnGlyProArgGluLysTyrGlyIleValAspTyrMet    260265270    IleGluGlnSerGlyProProSerLysGluIleLeuThrLeuLysGln    275280285    ValGlnGluPheLeuLysAspGlyAspAspValValIleIleGlyLeu    290295300    PheGlnGlyAspGlyAspProAlaTyrLeuGlnTyrGlnAspAlaAla    305310315320    AsnAsnLeuArgGluAspTyrLysPheHisHisThrPheSerProGlu    325330335    IleAlaLysPheLeuLysValSerLeuGlyLysLeuValLeuThrHis    340345350    ProGluLysPheGlnSerLysTyrGluProArgPheHisValMetAsp    355360365    ValGlnGlySerThrGluAlaSerAlaIleLysAspTyrValValLys    370375380    HisAlaLeuProLeuValGlyHisArgLysThrSerAsnAspAlaLys    385390395400    ArgTyrSerLysArgProLeuValValValTyrTyrSerValAspPhe    405410415    SerPheAspTyrArgAlaAlaThrGlnPheTrpArgAsnLysValLeu    420425430    GluValAlaLysAspPheProGluTyrThrPheAlaIleAlaAspGlu    435440445    GluAspTyrAlaThrGluValLysAspLeuGlyLeuSerGluSerGly    450455460    GluAspValAsnAlaAlaIleLeuAspGluSerGlyLysLysPheAla    465470475480    MetGluProGluGluPheAspSerAspThrLeuArgGluPheValThr    485490495    AlaPheLysLysGlyLysLeuLysProValIleLysSerGlnProVal    500505510    ProLysAsnAsnLysGlyProValLysValValValGlyLysThrPhe    515520525    AspAlaIleValMetAspProLysLysAspValLeuIleGluPheTyr    530535540    AlaProTrpCysGlyHisCysLysGlnLeuGluProIleTyrThrSer    545550555560    LeuGlyLysLysTyrLysGlyGlnLysAspLeuValIleAlaLysMet    565570575    AspAlaThrAlaAsnAspIleThrAsnAspGlnTyrLysValGluGly    580585590    PheProThrIleTyrPheAlaProSerGlyAspLysLysAsnProIle    595600605    LysPheGluGlyGlyAsnArgAspLeuGluHisLeuSerLysPheIle    610615620    AspGluHisAlaThrLysArgSerArgThrLysGluGluLeu    625630635    __________________________________________________________________________

I claim:
 1. An isolated active protein disulfide isomerase enzyme,comprising an amino acid sequence selected from the group consisting ofSEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ IDNO:10, and SEQ ID NO:
 12. 2. An enzyme composition comprising theprotein disulfide isomerase enzyme of claim
 1. 3. The composition ofclaim 2, in the form of a non-dusting granulate, stabilized liquid orprotected enzyme.
 4. The composition of claim 2, wherein the proteindisulfide isomerase is present in an amount between about 0.01-200 mg ofenzyme protein/g.
 5. The composition of claim 4, wherein the proteindisulfide isomerase is present in an amount between about 0.01-20 mg ofenzyme protein/g.
 6. The composition of claim 5, wherein the proteindisulfide isomerase is present in an amount between about 0.01-2 mg ofenzyme protein/g.
 7. The composition of claim 6, wherein the proteindisulfide isomerase is present in an amount between about 0.02-0.2 mg ofenzyme protein/g.
 8. The composition of claim 6, wherein the proteindisulfide isomerase is present in an amount between about 0.01-0.2 mg ofenzyme protein/g.
 9. The composition of claim 2 further comprisinganother enzyme selected from the group consisting of a protease, anamylase, a lipase, a peroxidase and a cellulase.
 10. A pharmaceuticalcomposition comprising the enzyme of claim
 1. 11. A process for treatingscleroproteins, comprising applying the composition of claim 2 to ascleroprotein.
 12. The process of claim 11, wherein said scleroproteinis human hair or skin, or animal hair or skin.
 13. The process of claim12, wherein said process involves waving, straightening, removing,degrading or softening of hair, or softening or restoration of skin. 14.The isolated enzyme of claim 1 obtained from Aspergillus oryzae, IFO4177 or Aspergillus niger 524 (ATCC 16882).
 15. An active proteindisulfide isomerase enzyme comprising the amino acid sequence of SEQ IDNO:11 or SEQ ID NO:
 4. 16. The isolated enzyme of claim 15 derived fromAspergillus oryzae, IFO 4177 or Aspergillus niger 524 (ATCC 16882). 17.An enzyme composition comprising the protein disulfide isomerase enzymeof claim
 15. 18. The composition of claim 17, in the form of anon-dusting granulate, stabilized liquid or protected enzyme.
 19. Thecomposition of claim 17, wherein the protein disulfide isomerase ispresent in an amount between about 0.01-200 mg of enzyme protein/g. 20.The composition of claim 19, wherein the protein disulfide isomerase ispresent in an amount between about 0.01-20 mg of enzyme protein/g. 21.The composition of claim 20, wherein the protein disulfide isomerase ispresent in an amount between about 0.01-2 mg of enzyme protein/g. 22.The composition of claim 21, wherein the protein disulfide isomerase ispresent in an amount between about 0.02-0.2 mg of enzyme protein/g. 23.The composition of claim 21, wherein the protein disulfide isomerase ispresent in an amount between about 0.01-0.2 mg of enzyme protein/g. 24.The composition of claim 17 further comprising another enzyme selectedfrom the group consisting of a protease, an amylase, a lipase, aperoxidase and a cellulase.
 25. A pharmaceutical composition comprisingthe enzyme of claim
 15. 26. A process for treating scleroproteins,comprising applying the composition of claim 17 to a scleroprotein. 27.The process of claim 26, wherein said scleroprotein is human hair orskin, or animal hair or skin.
 28. The process of claim 27, wherein saidprocess involves waving, straightening, removing, degrading or softeningof hair, or softening or restoration of skin.